Temporal Fidelity Research Articles

Multi-Scale Phenology of Temperate Grasslands: Improving Monitoring and Management With Near-Surface Phenocams

Grasslands of the Australian Southern Tablelands represent a patchwork of native and exotic systems, occupying a continuum of C3-dominated to C4-dominated grasslands where composition depends on disturbance factors (e.g. grazing) and climate. Managing these complex landscapes is both challenging and critical for maintaining the security of Australia’s pasture industries, and for protecting the biodiversity of native remnants. Differentiating C3 from C4 vegetation has been a prominent theme in remote sensing research due to distinct C3/C4 seasonal productivity patterns (phenology) and high uncertainty about how C3/C4 vegetation will respond to a changing climate. Phenology is used in northern hemisphere ecosystems for a range of purposes but has not been widely adopted in Australia, where dynamic climate often results in non-repetitive seasonal vegetation patterns. We employed time-lapse cameras (phenocams) to study the phenology of twelve grassland areas dominated by cool season (C3) and warm season (C4), native or exotic grasses near Canberra, Australia. Our aims were to assess phenological characteristics of the functional types and to determine the drivers of phenological variability. We compared the fine-scale phenocam seasonal profiles with field sampling and MODIS/Landsat satellite products to assess paddock-to-landscape functioning. We found C3/C4 species dominance to be the primary driver of phenological differences among grassland types, with C3 grasslands demonstrating peak greenness in spring, and senescing rapidly in response to high summer temperatures. In contrast, C4 grasslands showed peak activity in Austral summer and autumn (January-March). Some sites displayed primary and secondary peaks dependent on rainfall and species composition. We found that the proportion of dead vegetation is an important biophysical driver of grassland phenology, as were grazing pressures and species-dependent responses to rainfall and temperature. The satellite and field datasets were in general agreement with the phenocam results. However, the higher temporal fidelity of the cameras captured changes in vegetation not observed in the coarser satellite or field results. Our phenocam data shows consistent periods of increasing and decreasing greenness over as little as five days. Applications for management of grasslands in temperate Australia include the identification of remnant native grasslands, tracking biosecurity issues, and assessing productivity responses to climate variability.

Temporal Limits of Visual Motion Processing: Psychophysics and Neurophysiology.

Under optimal conditions, just 3–6 ms of visual stimulation suffices for humans to see motion. Motion perception on this timescale implies that the visual system under these conditions reliably encodes, transmits, and processes neural signals with near-millisecond precision. Motivated by in vitro evidence for high temporal precision of motion signals in the primate retina, we investigated how neuronal and perceptual limits of motion encoding relate. Specifically, we examined the correspondence between the time scale at which cat retinal ganglion cells in vivo represent motion information and temporal thresholds for human motion discrimination. The timescale for motion encoding by ganglion cells ranged from 4.6 to 91 ms, and depended non-linearly on temporal frequency, but not on contrast. Human psychophysics revealed that minimal stimulus durations required for perceiving motion direction were similarly brief, 5.6–65 ms, and similarly depended on temporal frequency but, above ~10%, not on contrast. Notably, physiological and psychophysical measurements corresponded closely throughout (r = 0.99), despite more than a 20-fold variation in both human thresholds and optimal timescales for motion encoding in the retina. The match in absolute values of the neurophysiological and psychophysical data may be taken to indicate that from the lateral geniculate nucleus (LGN) through to the level of perception little temporal precision is lost. However, we also show that integrating responses from multiple neurons can improve temporal resolution, and this potential trade-off between spatial and temporal resolution would allow for loss of temporal resolution after the LGN. While the extent of neuronal integration cannot be determined from either our human psychophysical or neurophysiological experiments and its contribution to the measured temporal resolution is unknown, our results demonstrate a striking similarity in stimulus dependence between the temporal fidelity established in the retina and the temporal limits of human motion discrimination.

Ratiometric fluorescence imaging for sodium selenite in living cells

Sodium selenite as a common selenium-containing agent has been extensively studied for human health, especially cancer prevention and treatment. However, precisely evaluating the dynamic changes of sodium selenite at cellular and molecular level in these pharmacological and pathophysiological studies is still challenging. Developing the chemical tools that enable the quantification of sodium selenite in vivo with high spatial and temporal fidelity thus can provide an efficient solution to this challenge. Here, we report the design, synthesis and biological evaluation of ratiometric sodium selenite fluorescent probe (HBTN-Se) for quantitative ratiometric fluorescence imaging of sodium selenite concentration fluctuations in living cells. A strategy for sodium selenite quantification has been achieved effectively through using a highly selective, rapid and biocompatible hydroxyl-deprotecting reaction. The ratiometric fluorescence imaging with HBTN-Se is constructed to precisely monitor changes in sodium selenite levels in living cells. Moreover, HBTN-Se is capable of visualizing differences in sodium selenite levels between normal cells and cancer cells, establishing the utility of this ratiometric detection platform for assessing the correlation between physiological effects of sodium selenite and its levels in living cells.

Generation of Monthly Precipitation Climatologies for Costa Rica Using Irregular Rain-Gauge Observational Networks

Precipitation climatologies for the period 1961–1990 were generated for all climatic regions of Costa Rica using an irregular rain-gauge observational network comprised by 416 rain-gauge stations. Two sub-networks were defined: a high temporal resolution sub-network (HTR), including stations having at least 20 years of continuous records during the study period (157 in total); and a high spatial resolution sub-network (HSR), which includes all HTR-stations plus those stations with less than 20 years of continuous records (416 in total). Results from the kriging variance reduction efficiency (KRE) objective function between the two sub-networks, show that ordinary kriging (OK) is unable to fully explain the spatio-temporal variability of precipitation within most climatic regions if only stations from the HTR sub-network are used. Results also suggests that in most cases, it is beneficial to increase the density of the rain-gauge observational network at the expense of temporal fidelity, by including more stations even though their records may not represent the same time step. Thereafter, precipitation climatologies were generated using seven deterministic (IDW, TS2, TS2PARA, TS2LINEAR, TPS, MQS and NN) and two geostatistical (OK and KED) interpolation methods. Performance of the various interpolation methods was evaluated using cross validation technique, selecting the mean absolute error (MAE) and the root-mean square error (RMSE) as agreement metrics. Results suggest that IDW is marginally superior to OK and KED for most climatic regions. The remaining deterministic methods however, considerably deviate from IDW, which suggests that these methods are incapable of properly capturing the true-nature of spatial precipitation patterns over the considered climatic regions. The final generated IDW climatology was then validated against the Global Precipitation Climatology Centre (GPCC), Climate Research Unit (CRU) and WorldClim datasets, in which overall spatial and temporal coherence is considered satisfactory, giving assurance about the use this new climatology in the development of local climate impact studies.

Calcium activation of cortical neurons by continuous electrical stimulation: Frequency dependence, temporal fidelity, and activation density.

Electrical stimulation of the brain has become a mainstay of fundamental neuroscience research and an increasingly prevalent clinical therapy. Despite decades of use in basic neuroscience research and the growing prevalence of neuromodulation therapies, gaps in knowledge regardingactivation or inactivation of neural elements over time have limited its ability to adequately interpret evoked downstream responses or fine-tune stimulation parameters to focus on desired responses. In this work,in vivotwo-photon microscopy was used to image neuronal calcium activity in layer 2/3 neurons of somatosensory cortex (S1) in male C57BL/6J-Tg(Thy1-GCaMP6s)GP4.3Dkim/J mice during 30s of continuous electrical stimulation at varying frequencies. We show frequency-dependent differences in spatial and temporal somatic responses during continuous stimulation. Our results elucidate conflicting results from prior studies reporting either dense spherical activationof somas biased toward those near the electrode, or sparse activation of somas at a distance via axons near the electrode. These findings indicate that the neural element specific temporal response local to the stimulating electrode changes as a function of applied charge density and frequency. These temporal responses need to be considered to properly interpret downstream circuit responses or determining mechanisms of action in basic science experiments or clinical therapeutic applications.

Optimized Chronos sets the clock for optogenetic hearing restoration.

The coming of age of optogenetics has motivated the development of clinical applications. Improved hearing restoration by optical cochlear implants is one such promising development. However, slow closing of light‐gated ion channels has remained an obstacle for achieving the high temporal fidelity required for optogenetic coding. In a study published in this issue, Keppeler et al demonstrate that optimized trafficking of the fast channelrhodopsin Chronos enables auditory nerve fibers to fire in response to light at near physiological rates.

Ultrafast optogenetic stimulation of the auditory pathway by targeting-optimized Chronos.

Optogenetic tools, providing non-invasive control over selected cells, have the potential to revolutionize sensory prostheses for humans. Optogenetic stimulation of spiral ganglion neurons (SGNs) in the ear provides a future alternative to electrical stimulation used in cochlear implants. However, most channelrhodopsins do not support the high temporal fidelity pertinent to auditory coding because they require milliseconds to close after light-off. Here, we biophysically characterized the fast channelrhodopsin Chronos and revealed a deactivation time constant of less than a millisecond at body temperature. In order to enhance neural expression, we improved its trafficking to the plasma membrane (Chronos-ES/TS). Following efficient transduction of SGNs using early postnatal injection of the adeno-associated virus AAV-PHPB into the mouse cochlea, fiber-based optical stimulation elicited optical auditory brainstem responses (oABR) with minimal latencies of 1ms, thresholds of 5μJ and 100μs per pulse, and sizable amplitudes even at 1,000Hz of stimulation. Recordings from single SGNs demonstrated good temporal precision of light-evoked spiking. In conclusion, efficient virus-mediated expression of targeting-optimized Chronos-ES/TS achieves ultrafast optogenetic control of neurons.

A decade of remotely sensed observations highlight complex processes linked to coastal permafrost bluff erosion in the Arctic

Eroding permafrost coasts are likely indicators and integrators of changes in the Arctic System as they are susceptible to the combined effects of declining sea ice extent, increases in open water duration, more frequent and impactful storms, sea-level rise, and warming permafrost. However, few observation sites in the Arctic have yet to link decadal-scale erosion rates with changing environmental conditions due to temporal data gaps. This study increases the temporal fidelity of coastal permafrost bluff observations using near-annual high spatial resolution (<1 m) satellite imagery acquired between 2008–2017 for a 9 km segment of coastline at Drew Point, Beaufort Sea coast, Alaska. Our results show that mean annual erosion for the 2007–2016 decade was 17.2 m yr−1, which is 2.5 times faster than historic rates, indicating that bluff erosion at this site is likely responding to changes in the Arctic System. In spite of a sustained increase in decadal-scale mean annual erosion rates, mean open water season erosion varied from 6.7 m yr−1 in 2010 to more than 22.0 m yr−1 in 2007, 2012, and 2016. This variability provided a range of coastal responses through which we explored the different roles of potential environmental drivers. The lack of significant correlations between mean open water season erosion and the environmental variables compiled in this study indicates that we may not be adequately capturing the environmental forcing factors, that the system is conditioned by long-term transient effects or extreme weather events rather than annual variability, or that other not yet considered factors may be responsible for the increased erosion occurring at Drew Point. Our results highlight an increase in erosion at Drew Point in the 21st century as well as the complexities associated with unraveling the factors responsible for changing coastal permafrost bluffs in the Arctic.

Ca2+ Regulates the Kinetics of Synaptic Vesicle Fusion at the Afferent Inner Hair Cell Synapse.

The early auditory pathway processes information at high rates and with utmost temporal fidelity. Consequently, the synapses in the auditory pathway are highly specialized to meet the extraordinary requirements on signal transmission. The calyceal synapses in the auditory brainstem feature more than a hundred active zones (AZs) with thousands of releasable synaptic vesicles (SVs). In contrast, the first auditory synapse, the afferent synapse of inner hair cells (IHCs) and type I spiral ganglion neurons (SGNs), typically exhibits a single ribbon-type AZ tethering only tens of SVs resulting in a highly stochastic pattern of transmitter release. Spontaneous excitatory postsynaptic currents (sEPSCs), besides more conventional EPSCs with a single peak, fast rise and decay (compact), also include EPSCs with multiple peaks, variable rise and decay times (non-compact). The strong heterogeneity in size and shape of spontaneous EPSCs has led to the hypothesis of multivesicular release (MVR) that is more (compact) or less (non-compact) synchronized by coordination of release sites. Alternatively, univesicular release (UVR), potentially involving glutamate release through a flickering fusion pore for non-compact EPSCs, has been suggested to underlie IHC exocytosis. Here, we further investigated the mode of release by recording sEPSCs from SGNs of hearing rats while manipulating presynaptic IHC Ca2+ influx by changes in extracellular [Ca2+] ([Ca2+]e) and by application of the Ca2+ channel antagonist, isradipine, or the Ca2+ channel agonist, BayK8644 (BayK). Our data reveal that Ca2+ influx manipulation leaves the distributions of sEPSC amplitude and charge largely unchanged. Regardless the type of manipulation, the rate of sEPSC decreased with the reduction in Ca2+ influx. The fraction of compact sEPSCs was increased in the presence of BayK, an effect that was abolished when combined with decreased [Ca2+]e. In conclusion, we propose that UVR is the prevailing mode of exocytosis at cochlear IHCs of hearing rats, whereby the rate of exocytosis and the kinetics of SV fusion are regulated by Ca2+ influx.

Balancing presynaptic release and endocytic membrane retrieval at hair cell ribbon synapses.

The timely and reliable processing of auditory and vestibular information within the inner ear requires highly sophisticated sensory transduction pathways. On a cellular level, these demands are met by hair cells, which respond to sound waves - or alterations in body positioning - by releasing glutamate-filled synaptic vesicles (SVs) from their presynaptic active zones with unprecedented speed and exquisite temporal fidelity, thereby initiating the auditory and vestibular pathways. In order to achieve this, hair cells have developed anatomical and molecular specializations, such as the characteristic and name-giving 'synaptic ribbons' - presynaptically anchored dense bodies that tether SVs prior to release - as well as other unique or unconventional synaptic proteins. The tightly orchestrated interplay between these molecular components enables not only ultrafast exocytosis, but similarly rapid and efficient compensatory endocytosis. So far, the knowledge of how endocytosis operates at hair cell ribbon synapses is limited. In this Review, we summarize recent advances in our understanding of the SV cycle and molecular anatomy of hair cell ribbon synapses, with a focus on cochlear inner hair cells.

Innovation in rangeland monitoring: annual, 30 m, plant functional type percent cover maps for U.S. rangelands, 1984–2017

AbstractInnovations in machine learning and cloud‐based computing were merged with historical remote sensing and field data to provide the first moderate resolution, annual, percent cover maps of plant functional types across rangeland ecosystems to effectively and efficiently respond to pressing challenges facing conservation of biodiversity and ecosystem services. We utilized the historical Landsat satellite record, gridded meteorology, abiotic land surface data, and over 30,000 field plots within a Random Forests model to predict per‐pixel percent cover of annual forbs and grasses, perennial forbs and grasses, shrubs, and bare ground over the western United States from 1984 to 2017. Results were validated using three independent collections of plot‐level measurements, and resulting maps display land cover variation in response to changes in climate, disturbance, and management. The maps, which will be updated annually at the end of each year, provide exciting opportunities to expand and improve rangeland conservation, monitoring, and management. The data open new doors for scientific investigation at an unprecedented blend of temporal fidelity, spatial resolution, and geographic scale.

Long-term all-optical interrogation of cortical neurons in awake-behaving nonhuman primates.

Whereas optogenetic techniques have proven successful in their ability to manipulate neuronal populations—with high spatial and temporal fidelity—in species ranging from insects to rodents, significant obstacles remain in their application to nonhuman primates (NHPs). Robust optogenetics-activated behavior and long-term monitoring of target neurons have been challenging in NHPs. Here, we present a method for all-optical interrogation (AOI), integrating optical stimulation and simultaneous two-photon (2P) imaging of neuronal populations in the primary visual cortex (V1) of awake rhesus macaques. A red-shifted channel-rhodopsin transgene (ChR1/VChR1 [C1V1]) and genetically encoded calcium indicators (genetically encoded calmodulin protein [GCaMP]5 or GCaMP6s) were delivered by adeno-associated viruses (AAVs) and subsequently expressed in V1 neuronal populations for months. We achieved optogenetic stimulation using both single-photon (1P) activation of neuronal populations and 2P activation of single cells, while simultaneously recording 2P calcium imaging in awake NHPs. Optogenetic manipulations of V1 neuronal populations produced reliable artificial visual percepts. Together, our advances show the feasibility of precise and stable AOI of cortical neurons in awake NHPs, which may lead to broad applications in high-level cognition and preclinical testing studies.

Incoherent clocking in coherent radio interferometers

Radio interferometer arrays are instruments requiring real-time synchronisation of all array elements to capture and align the observed source wavefront prior to further processing such as correlation and/or beamforming. One method is to deliver a frequency, from a common central local oscillator (LO), using a round-trip phase-stable distribution system such that each element has its own phase-stable copy of the clock with required real-time fidelity for down-conversion and digitisation. The method described here involves equipping each array element with its own independent ‘eLO’ and sending a ‘tracer’ of it to a central site, where the difference between the elements’ eLOs and the central ‘cLO’ is measured. The digitised data from each element is corrected at the central site according to the measurement, before correlation/beamforming. The advantage of this method is that only a ‘trace’ of eLO, needed to measure the rate of change of eLO relative to cLO integrated over a measurement interval, needs to be conveyed over this tracer link, rather than the frequency itself, thereby requiring lower link temporal fidelity and bandwidth.

Climatic Drivers of Ponderosa Pine Growth in Central Idaho

Despite the widespread use of ponderosa pine as an important hydroclimate proxy, we actually understand very little about its climate response in the Northern Rockies. Here, we analyze two new ponderosa pine chronologies to investigate how climate influences annual growth. Despite differences in precipitation amount and timing and large elevation differences (1820 m versus 1060 m), ring width at both sites was strongly driven by water availability. The mid-elevation, water-limited site responded well to previous fall precipitation whereas the wetter, high-elevation site responded to growing season precipitation and temperature. When precipitation and temperature were simultaneously accounted for using the standardized precipitation evapotranspiration index, ring-width response between sites converged and appeared nearly identical. Water stress drove the timing of ponderosa pine growth by a combination of factors such as strong water dependence, and determinate growth physiology, as indicated by lag-1 autocorrelation. When analyzing response to single-month climate variables, precipitation from growing-season months dominates. When we examined seasonal variables, climate from the previous year became more important. Temporal fidelity of the climatic response at both sites maintained significance across the historical record, although the relationship weakened at the low-elevation site. The collection of new tree-ring data sets such as these for central Idaho improves our understanding of ponderosa pine growth response to climate.

Geometric classification of brain network dynamics via conic derivative discriminants

BackgroundOver the past decade, pattern decoding techniques have granted neuroscientists improved anatomical specificity in mapping neural representations associated with function and cognition. Dynamical patterns are of particular interest, as evidenced by the proliferation and success of frequency domain methods that reveal structured spatiotemporal rhythmic brain activity. One drawback of such approaches, however, is the need to estimate spectral power, which limits the temporal resolution of classification. New methodWe propose an alternative method that enables classification of dynamical patterns with high temporal fidelity. The key feature of the method is a conversion of time-series data into temporal derivatives. By doing so, dynamically-coded information may be revealed in terms of geometric patterns in the phase space of the derivative signal. ResultsWe derive a geometric classifier for this problem which simplifies into a straightforward calculation in terms of covariances. We demonstrate the relative advantages and disadvantages of the technique with simulated data and benchmark its performance with an EEG dataset of covert spatial attention. We reveal the timecourse of covert spatial attention and, by mapping the classifier weights anatomically, its retinotopic organization. Comparison with existing methodWe especially highlight the ability of the method to provide strong group-level classification performance compared to existing benchmarks, while providing information that is complementary with classical spectral-based techniques. The robustness and sensitivity of the method to noise is also examined relative to spectral-based techniques. ConclusionThe proposed classification technique enables decoding of dynamic patterns with high temporal resolution, performs favorably to benchmark methods, and facilitates anatomical inference.

Unsupervised Discovery of Demixed, Low-Dimensional Neural Dynamics across Multiple Timescales through Tensor Component Analysis

Perceptions, thoughts, and actions unfold over millisecond timescales, while learned behaviors can require many days to mature. While recent experimental advances enable large-scale and long-term neural recordings with high temporal fidelity, it remains a formidable challenge to extract unbiased and interpretable descriptions of how rapid single-trial circuit dynamics change slowly over many trials to mediate learning. We demonstrate a simple tensor component analysis (TCA) can meet this challenge by extracting three interconnected, low-dimensional descriptions of neural data: neuron factors, reflecting cell assemblies; temporal factors, reflecting rapid circuit dynamics mediating perceptions, thoughts, and actions within each trial; and trial factors, describing both long-term learning and trial-to-trial changes in cognitive state. We demonstrate the broad applicability of TCA by revealing insights intodiverse datasets derived from artificial neural networks, large-scale calcium imaging of rodent prefrontal cortex during maze navigation, and multielectrode recordings of macaque motor cortex during brain machine interface learning.

Ecophenotypy, temporal and spatial fidelity, functional morphology, and physiological trade-offs among intertidal bivalves

AbstractEcophenotypic variation in populations is driven by differences in environmental variables. In marine environments, ecophenotypic variation may be caused by differences in hydrodynamic conditions, substrate type, water depth, temperature, salinity, oxygen concentration, and habitat heterogeneity, among others. Instances of ecophenotypic variation in modern and fossil settings are common, but little is known about the influences of time averaging and spatial averaging on their preservation. Here we examine the shell morphology of two adjacent populations, both live collected and death assemblages, of the infaunal, suspension-feeding, intertidal bivalve Leukoma staminea from the well-studied Argyle Creek and Argyle Lagoon locations on San Juan Island, Washington. Individuals in the low-energy lagoon are free to burrow in the fine-grained substrate, while clams in the high-energy creek are precluded from burrowing in the rocky channel. Our results demonstrate variation in size and shape between the adjacent habitats. Lagoon clams are larger, more disk-shaped, and have relatively larger siphons than their creek counterparts, which are smaller, more spherical in shape, and have a relatively shallower pallial sinus. This ecophenotypy is preserved among death assemblages, although with generally greater variation due to time averaging and shell transport. Our interpretation is that ecophenotypic variation, in this case, is induced by differing hydrodynamic regimes and substrate types, cumulatively resulting in physiological trade-offs diverting resources from feeding and respiration to stability and shell strength, all of which have the potential to be preserved in the fossil record.

The effects of aging on auditory cortical function

Age-related hearing loss is a prominent deficit, afflicting approximately half of the geriatric population. In many cases, the person may have no deficits in detecting sounds, but nonetheless suffers from a reduced ability to understand speech, particularly in a noisy environment. While rodent models have shown that there are a variety of age-related changes throughout the auditory neuraxis, far fewer studies have investigated the effects at the cortical level. Here I review recent evidence from a non-human primate model of age-related hearing loss at the level of the core (primary auditory cortex, A1) and belt (caudolateral field, CL) in young and aged animals with normal detection thresholds. The findings are that there is an increase in both the spontaneous and driven activity, an increase in spatial tuning, and a reduction in the temporal fidelity of the response in aged animals. These results are consistent with an age-related imbalance of excitation and inhibition in the auditory cortex. These spatial and temporal processing deficits could underlie the major complaint of geriatrics, that it is difficult to understand speech in noise.

High frequency neural spiking and auditory signaling by ultrafast red-shifted optogenetics

Optogenetics revolutionizes basic research in neuroscience and cell biology and bears potential for medical applications. We develop mutants leading to a unifying concept for the construction of various channelrhodopsins with fast closing kinetics. Due to different absorption maxima these channelrhodopsins allow fast neural photoactivation over the whole range of the visible spectrum. We focus our functional analysis on the fast-switching, red light-activated Chrimson variants, because red light has lower light scattering and marginal phototoxicity in tissues. We show paradigmatically for neurons of the cerebral cortex and the auditory nerve that the fast Chrimson mutants enable neural stimulation with firing frequencies of several hundred Hz. They drive spiking at high rates and temporal fidelity with low thresholds for stimulus intensity and duration. Optical cochlear implants restore auditory nerve activity in deaf mice. This demonstrates that the mutants facilitate neuroscience research and future medical applications such as hearing restoration.

Cholinergic innervation of principal neurons in the cochlear nucleus of the Mongolian gerbil.

Principal neurons in the ventral cochlear nucleus (VCN) receive powerful ascending excitation and pass on the auditory information with exquisite temporal fidelity. Despite being dominated by ascending inputs, the VCN also receives descending cholinergic connections from olivocochlear neurons and from higher regions in the pontomesencephalic tegmentum. In Mongolian gerbils, acetylcholine acts as an excitatory and modulatory neurotransmitter on VCN neurons, but the anatomical structure of cholinergic innervation of gerbil VCN is not well described. We applied fluorescent immunohistochemical staining to elucidate the development and the cellular localization of presynaptic and postsynaptic components of the cholinergic system in the VCN of the Mongolian gerbil. We found that cholinergic fibers (stained with antibodies against the vesicular acetylcholine transporter) were present before hearing onset at P5, but innervation density increased in animals after P10. Early in development cholinergic fibers invaded the VCN from the medial side, spread along the perimeter and finally innervated all parts of the nucleus only after the onset of hearing. Cholinergic fibers ran in a rostro-caudal direction within the nucleus and formed en-passant swellings in the neuropil between principal neurons. Nicotinic and muscarinic receptors were expressed differentially in the VCN, with nicotinic receptors being mostly expressed in dendritic areas while muscarinic receptors were located predominantly in somatic membranes. These anatomical data support physiological indications that cholinergic innervation plays a role in modulating information processing in the cochlear nucleus.