Characterization of cerebrospinal fluid biomarkers associated with neurodegenerative diseases in healthy cynomolgus and rhesus macaque monkeys.

Monkeys are becoming important translational models of neurodegenerative disease. To facilitate model development, we measured cerebrospinal fluid (CSF) concentrations of key biomarkers in healthy male and female cynomolgus and rhesus macaques. Amyloid beta (Aß40, Aß42), tau (total tau [t-tau], phosphorylated tau [pThr181]), and neurofilament light (NfL) concentrations were measured in CSF of 82 laboratory-housed, experimentally naïve cynomolgus (n = 33) and rhesus (n = 49) macaques. Aß40 and Aß42 were significantly higher in rhesus, and female rhesus were higher than males. NfL and t-tau were higher in males, and NfL was higher in rhesus macaques. p-tau was not affected by species or sex. We also examined whether sample location (lumbar or cisterna puncture) affected concentrations. Sample acquisition site only affected NfL, which was higher in CSF from lumbar puncture compared to cisterna magna puncture. Establishing normative biomarker values for laboratory-housed macaque monkeys provides an important resource by which to compare to monkey models of neurodegenerative diseases.

The effect of lumbar puncture on the neurodegeneration biomarker neurofilament light in macaque monkeys.

INTRODUCTION: Neurofilament light (NFL) in cerebrospinal fluid (CSF) is elevated in neurodegenerative disease patients, and may track disease progression and treatment. Macaque monkeys are emerging as important translational models of neurodegeneration, and NFL may be a useful biomarker. METHODS: To determine the influence of a previous lumbar puncture (LP) on NFL, we collected CSF at multiple time points in macaque monkeys via LP or cisterna magna puncture. NFL, amyloid beta (Aß40, Aß42), and tau (tTau, pTau) in CSF were measured by standard enzyme-linked immunosorbent assay and multiplex. RESULTS: NFL was significantly elevated at 14 to 23 days after an LP (median increase: 162%). Aß and tau biomarkers remained stable. NFL peaked and decayed over 1 to 2 months after LP. NFL was not elevated after cisterna magna puncture. DISCUSSION: Results suggest damage of the cauda equina during LP may increase NFL. Caution should be taken in interpreting NFL concentration in studies in which repeat LPs are performed.

Behavioral shaping of rhesus macaques using the Cambridge neuropsychological automated testing battery.

BACKGROUND: The Cambridge neuropsychological test automated battery (CANTAB) is a set of computerized visuospatial tests used to probe cognition in humans. The non-human primate (NHP) version of the battery is a valuable translational research tool to quantify cognitive changes in NHP models of disease by allowing direct comparison with performance data from human patient populations. One limitation is the long training times required for NHPs to reach appropriate levels of task performance, which is prohibitive for high throughput experimental designs. NEW METHOD: We report a new training regimen to teach NHPs a subset of CANTAB cognitive tasks using a method of successive approximations (shaping), where rewarded behaviors progressively approximate the goal behavior, and sequential task learning is used to build upon previously learned rules. Using this refined method, we taught 9 adult rhesus macaques to perform three tasks: the self-ordered spatial search (SOSS), delayed match-to-sample (DMTS), and paired associative learning (PAL) tasks. RESULTS AND COMPARISON WITH EXISTING METHODS: NHPs learned all three cognitive tasks in approximately 130 training sessions, roughly 200 sessions faster than previously published training times. NHPs were able to perform each task to a stable level of performance (>80 % correct) enabling their use in future cognitive experiments. CONCLUSIONS: Our approach of behavioral shaping reduced the time to train NHPs to performance criteria on SOSS, DMTS, and PAL tasks. This allows efficient use of the NHP-adapted CANTAB to compare cognitive changes in NHP models of neurological disease with those observed in human patient populations.

Understanding the link between insulin resistance and Alzheimer's disease: Insights from animal models.

Alzheimer's disease (AD) is a devastating neurodegenerative disease affecting millions of people worldwide. AD is characterized by a profound impairment of higher cognitive functions and still lacks any effective disease-modifying treatment. Defective insulin signaling has been implicated in AD pathophysiology, but the mechanisms underlying this process are not fully understood. Here, we review the molecular mechanisms underlying defective brain insulin signaling in rodent models of AD, and in a non-human primate (NHP) model of the disease that recapitulates features observed in AD brains. We further highlight similarities between the NHP and human brains and discuss why NHP models of AD are important to understand disease mechanisms and to improve the translation of effective therapies to humans. We discuss how studies using different animal models have contributed to elucidate the link between insulin resistance and AD.

The diabetes drug liraglutide reverses cognitive impairment in mice and attenuates insulin receptor and synaptic pathology in a non-human primate model of Alzheimer's disease.

Alzheimer's disease (AD) is a devastating neurological disorder that still lacks an effective treatment, and this has stimulated an intense pursuit of disease-modifying therapeutics. Given the increasingly recognized link between AD and defective brain insulin signaling, we investigated the actions of liraglutide, a glucagon-like peptide-1 (GLP-1) analog marketed for treatment of type 2 diabetes, in experimental models of AD. Insulin receptor pathology is an important feature of AD brains that impairs the neuroprotective actions of central insulin signaling. Here, we show that liraglutide prevented the loss of brain insulin receptors and synapses, and reversed memory impairment induced by AD-linked amyloid-ß oligomers (AßOs) in mice. Using hippocampal neuronal cultures, we determined that the mechanism of neuroprotection by liraglutide involves activation of the PKA signaling pathway. Infusion of AßOs into the lateral cerebral ventricle of non-human primates (NHPs) led to marked loss of insulin receptors and synapses in brain regions related to memory. Systemic treatment of NHPs with liraglutide provided partial protection, decreasing AD-related insulin receptor, synaptic, and tau pathology in specific brain regions. Synapse damage and elimination are amongst the earliest known pathological changes and the best correlates of memory impairment in AD. The results illuminate mechanisms of neuroprotection by liraglutide, and indicate that GLP-1 receptor activation may be harnessed to protect brain insulin receptors and synapses in AD. © 2018 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

Multisensory integration in orienting behavior: Pupil size, microsaccades, and saccades.

Signals from different sensory modalities are integrated in the brain to optimize behavior. Although multisensory integration has been demonstrated in saccadic eye movements, its influence on other orienting responses, including pupil size and microsaccades, is still poorly understood. We examined human gaze orienting responses following presentation of visual, auditory, or combined audiovisual stimuli. Transient pupil dilation and microsaccade inhibition were evoked shortly after the appearance of a salient stimulus. Audiovisual stimuli evoked larger pupil dilation, greater microsaccade inhibition, and faster saccade reaction times compared to unimodal conditions. Trials with faster saccadic reaction times were accompanied with greater pupil dilation responses. Similar modulation of pre-stimulus pupil-size-change rate was observed between stimulus-evoked saccadic and pupillary responses. Thus, multisensory integration impacts multiple components of orienting, with coordination between saccade and pupil responses, implicating the superior colliculus in coordinating these responses because of its central role in both orienting behavior and multisensory integration.

Spatio-temporal response properties of local field potentials in the primate superior colliculus.

Local field potentials (LFPs) are becoming increasingly popular in neurophysiological studies. However, to date, most of the knowledge about LFPs has been obtained from cortical recordings. Here, we recorded single unit activity (SUA) and LFPs simultaneously from the superior colliculus (SC) of behaving rhesus monkeys. The SC is a midbrain structure that plays a central role in the visual orienting response. Previous studies have characterised the visual and visuomotor response properties of SUA in the superficial layers of the SC and the intermediate layers of the SC, respectively. We found that the signal properties of SUA were well preserved in the LFPs recorded from the SC. The SUA and LFPs had similar spatial and temporal properties, and the response properties of LFPs differed across layers, i.e. purely visual in the superficial layers of the SC but showing significant motor responses in the intermediate layers of the SC. There were also differences between SUA and LFPs. LFPs showed a significant reversal of activity following the phasic visual response, suggesting that the neighboring neurons were suppressed. The results indicate that the LFP can be used as a reliable measure of the SC activity in lieu of SUA, and open up a new way to assess sensorimotor processing within the SC.

Alzheimer's disease-like pathology induced by amyloid-ß oligomers in nonhuman primates.

Alzheimer's disease (AD) is a devastating neurodegenerative disorder and a major medical problem. Here, we have investigated the impact of amyloid-ß (Aß) oligomers, AD-related neurotoxins, in the brains of rats and adult nonhuman primates (cynomolgus macaques). Soluble Aß oligomers are known to accumulate in the brains of AD patients and correlate with disease-associated cognitive dysfunction. When injected into the lateral ventricle of rats and macaques, Aß oligomers diffused into the brain and accumulated in several regions associated with memory and cognitive functions. Cardinal features of AD pathology, including synapse loss, tau hyperphosphorylation, astrocyte and microglial activation, were observed in regions of the macaque brain where Aß oligomers were abundantly detected. Most importantly, oligomer injections induced AD-type neurofibrillary tangle formation in the macaque brain. These outcomes were specifically associated with Aß oligomers, as fibrillar amyloid deposits were not detected in oligomer-injected brains. Human and macaque brains share significant similarities in terms of overall architecture and functional networks. Thus, generation of a macaque model of AD that links Aß oligomers to tau and synaptic pathology has the potential to greatly advance our understanding of mechanisms centrally implicated in AD pathogenesis. Furthermore, development of disease-modifying therapeutics for AD has been hampered by the difficulty in translating therapies that work in rodents to humans. This new approach may be a highly relevant nonhuman primate model for testing therapeutic interventions for AD.

Transient pupil response is modulated by contrast-based saliency.

The sudden appearance of a novel stimulus in the environment initiates a series of orienting responses that include coordinated shifts of gaze and attention, and also transient changes in pupil size. Although numerous studies have identified a significant effect of stimulus saliency on shifts of gaze and attention, saliency effects on pupil size are less understood. To examine salience-evoked pupil responses, we presented visual, auditory, or audiovisual stimuli while monkeys fixated a central visual spot. Transient pupil dilation was elicited after visual stimulus presentation regardless of target luminance relative to background, and auditory stimuli also evoked similar pupil responses. Importantly, the evoked pupil response was modulated by contrast-based saliency, with faster and larger pupil responses following the presentation of more salient stimuli. The initial transient component of pupil dilation was qualitatively similar to that evoked by weak microstimulation of the midbrain superior colliculus. The pupil responses elicited by audiovisual stimuli were well predicted by a linear summation of each modality response. Together, the results suggest that the transient pupil response, as one component of orienting, is modulated by contrast-based saliency, and the superior colliculus is likely involved in its coordination.

Competitive integration of visual and goal-related signals on neuronal accumulation rate: a correlate of oculomotor capture in the superior colliculus.

The mechanisms that underlie the integration of visual and goal-related signals for the production of saccades remain poorly understood. Here, we examined how spatial proximity of competing stimuli shapes goal-directed responses in the superior colliculus (SC), a midbrain structure closely associated with the control of visual attention and eye movements. Monkeys were trained to perform an oculomotor-capture task [Theeuwes, J., Kramer, A. F., Hahn, S., Irwin, D. E., & Zelinsky, G. J. Influence of attentional capture on oculomotor control. Journal of Experimental Psychology. Human Perception and Performance, 25, 1595-1608, 1999], in which a target singleton was revealed via an isoluminant color change in all but one item. On a portion of the trials, an additional salient item abruptly appeared near or far from the target. We quantified how spatial proximity between the abrupt-onset and the target shaped the goal-directed response. We found that the appearance of an abrupt-onset near the target induced a transient decrease in goal-directed discharge of SC visuomotor neurons. Although this was indicative of spatial competition, it was immediately followed by a rebound in presaccadic activation, which facilitated the saccadic response (i.e., it induced shorter saccadic RT). A similar suppression also occurred at most nontarget locations even in the absence of the abrupt-onset. This is indicative of a mechanism that enabled monkeys to quickly discount stimuli that shared the common nontarget feature. These results reveal a pattern of excitation/inhibition across the SC visuomotor map that acted to facilitate optimal behavior-the short duration suppression minimized the probability of capture by salient distractors, whereas a subsequent boost in accumulation rate ensured a fast goal-directed response. Such nonlinear dynamics should be incorporated into future biologically plausible models of saccade behavior.

Microstimulation of the monkey superior colliculus induces pupil dilation without evoking saccades.

The orienting reflex is initiated by a salient stimulus and facilitates quick, appropriate action. It involves a rapid shift of the eyes, head, and attention and other physiological responses such as changes in heart rate and transient pupil dilation. The SC is a critical structure in the midbrain that selects incoming stimuli based on saliency and relevance to coordinate orienting behaviors, particularly gaze shifts, but its causal role in pupil dilation remains poorly understood in mammals. Here, we examined the role of the primate SC in the control of pupil dynamics. While requiring monkeys to keep their gaze fixed, we delivered weak electrical microstimulation to the SC, so that saccadic eye movements were not evoked. Pupil size increased transiently after microstimulation of the intermediate SC layers (SCi) and the size of evoked pupil dilation was larger on a dim versus bright background. In contrast, microstimulation of the superficial SC layers did not cause pupil dilation. Thus, the SCi is directly involved not only in shifts of gaze and attention, but also in pupil dilation as part of the orienting reflex, and the function of pupil dilation may be related to increasing visual sensitivity. The shared neural mechanisms suggest that pupil dilation may be associated with covert attention.

Visual adaptation and novelty responses in the superior colliculus.

The brain's ability to ignore repeating, often redundant, information while enhancing novel information processing is paramount to survival. When stimuli are repeatedly presented, the response of visually sensitive neurons decreases in magnitude, that is, neurons adapt or habituate, although the mechanism is not yet known. We monitored the activity of visual neurons in the superior colliculus (SC) of rhesus monkeys who actively fixated while repeated visual events were presented. We dissociated adaptation from habituation as mechanisms of the response decrement by using a Bayesian model of adaptation, and by employing a paradigm including rare trials that included an oddball stimulus that was either brighter or dimmer. If the mechanism is adaptation, response recovery should be seen only for the brighter stimulus; if the mechanism is habituation, response recovery ('dishabituation') should be seen for both the brighter and dimmer stimuli. We observed a reduction in the magnitude of the initial transient response and an increase in response onset latency with stimulus repetition for all visually responsive neurons in the SC. Response decrement was successfully captured by the adaptation model, which also predicted the effects of presentation rate and rare luminance changes. However, in a subset of neurons with sustained activity in response to visual stimuli, a novelty signal akin to dishabituation was observed late in the visual response profile for both brighter and dimmer stimuli, and was not captured by the model. This suggests that SC neurons integrate both rapidly discounted information about repeating stimuli and novelty information about oddball events, to support efficient selection in a cluttered dynamic world.

The eccentricity effect for auditory saccadic reaction times is independent of target frequency.

Although much is understood about the stimulus properties affecting the latency of saccadic eye movements to visual targets, relatively little is known about the properties affecting saccades to auditory targets. This study examined the effect of three primary acoustic features-frequency, intensity, and spatial location-on auditory saccade characteristics in humans, and compared them to visual saccades. Saccade targets were presented from an azimuthal array of speakers and LEDs spanning +/-36 degrees. There was an 'eccentricity effect' for auditory saccades such that latencies decreased by up to 70 ms with eccentricity. This was observed for all frequencies and intensities tested. There was a smaller effect in the opposite direction effect for visual saccades. Auditory saccades had similar latencies to visual saccades (within 5 ms) for near midline locations, but were up to 90 ms faster at eccentric locations (+/-36 degrees). Overall, saccadic latencies were shortest for wideband noise and narrowband noises with center frequencies falling within the human speech range. Examination of saccade accuracy showed decreasing accuracy with increasing eccentricity, and a negative correlation between accuracy and latency for auditory stimuli.

Color-related signals in the primate superior colliculus.

Color is important for segmenting objects from backgrounds, which can in turn facilitate visual search in complex scenes. However, brain areas involved in orienting the eyes toward colored stimuli in our environment are not believed to have access to color information. Here, we show that neurons in the intermediate layers of the monkey superior colliculus (SC), a critical structure for the production of saccadic eye movements, can respond to isoluminant color stimuli with the same magnitude as a maximum contrast luminance stimulus. In contrast, neurons from the superficial SC layers showed little color-related activity. Crucially, visual onset latencies were 30-35 ms longer for color, implying that luminance and chrominance information reach the SC through distinct pathways and that the observed color-related activity is not the result of residual luminance signals. Furthermore, these differences in visual onset latency translated directly into differences in saccadic reaction time. The results demonstrate that the saccadic system can signal the presence of chromatic stimuli only one stage from the brainstem premotor circuitry that drives the eyes.

Free viewing of dynamic stimuli by humans and monkeys.

Due to extensive homologies, monkeys provide a sophisticated animal model of human visual attention. However, for electrophysiological recording in behaving animals simplified stimuli and controlled eye position are traditionally used. To validate monkeys as a model for human attention during realistic free viewing, we contrasted human (n = 5) and monkey (n = 5) gaze behavior using 115 natural and artificial video clips. Monkeys exhibited broader ranges of saccadic endpoints and amplitudes and showed differences in fixation and intersaccadic intervals. We compared tendencies of both species to gaze toward scene elements with similar low-level visual attributes using two computational models--luminance contrast and saliency. Saliency was more predictive of both human and monkey gaze, predicting human saccades better than monkey saccades overall. Quantifying interobserver gaze consistency revealed that while humans were highly consistent, monkeys were more heterogeneous and were best predicted by the saliency model. To address these discrepancies, we further analyzed high-interest gaze targets--those locations simultaneously chosen by at least two monkeys. These were on average very similar to human gaze targets, both in terms of specific locations and saliency values. Although substantial quantitative differences were revealed, strong similarities existed between both species, especially when focusing analysis onto high-interest targets.

On the importance of the transient visual response in the superior colliculus.

A salient event in the environment can initiate a complex orienting response that includes a shift in gaze. The midbrain superior colliculus (SC) contains the appropriate circuitry to generate and distribute a signal of the priority of this event, and co-ordinate the orienting response. The magnitude and timing of the short-latency transient visual response in the SC, when combined with cortical inputs signaling stimulus relevance and expectation, influences the type and latency of the orienting response. This signal in the SC is distributed to higher cortical areas to influence visual processing, to the reinforcement learning system to influence future actions, and to premotor circuits, including neck and shoulder muscles, to influence immediate action.

Cue repetition increases inhibition of return.

Inhibition of return (IOR) refers to slowed responses to targets presented at the same location as a preceding stimulus. We explored whether the IOR effect would increase with the number of cues preceding the target (a 'cue'). Subjects performed a Posner cueing task with 1-5 cue presentations prior to the target, to which they made either a manual localization (Experiment 1) or target discrimination response (Experiment 2). The cues could be the same as (Experiment 1), or differ in shape from (Experiment 2), the target. The results showed that regardless of cue-target congruency the IOR effect increased dramatically with the number of preceding cues. This increase was driven mostly by a linear slowing of reaction times to targets presented on the same side as the cue(s), suggesting that a process such as sensory adaptation and/or habituation may be a contributing mechanism to the IOR effect.

Physiological evidence that pyramidal neurons lack functional water channels.

The physiological conditions that swell mammalian neurons are clinically important but contentious. Distinguishing the neuronal component of brain swelling requires viewing intact neuronal cell bodies, dendrites, and axons and measuring their changing volume in real time. Cultured or dissociated neuronal somata swell within minutes under acutely overhydrated conditions and shrink when strongly dehydrated. But paradoxically, most central nervous system (CNS) neurons do not express aquaporins, the membrane channels that conduct osmotically driven water. Using 2-photon laser scanning microscopy (2PLSM), we monitored neuronal volume under osmotic stress in real time. Specifically, the volume of pyramidal neurons in cerebral cortex and axon terminals comprising cerebellar mossy fibers was measured deep within live brain slices. The expected swelling or shrinking of the gray matter was confirmed by recording altered light transmittance and by indirectly measuring extracellular resistance over a wide osmotic range of -80 to +80 milliOsmoles (mOsm). Neurons expressing green fluorescent protein were then imaged with 2PLSM between -40 and +80 mOsm over 20 min. Surprisingly, pyramidal somata, dendrites, and spines steadfastly maintained their volume, as did the cerebellar axon terminals. This precluded a need for the neurons to acutely regulate volume, preserved their intrinsic electrophysiological stability, and confirmed that these CNS nerve cells lack functional aquaporins. Thus, whereas water easily permeates the aquaporin-rich endothelia and glia driving osmotic brain swelling, neurons tenatiously maintain their volume. However, these same neurons then swell dramatically upon oxygen/glucose deprivation or [K+]0 elevation, so prolonged depolarization (as during stroke or seizure) apparently swells neurons by opening nonaquaporin channels to water.

Auditory saltation in the vertical midsagittal plane.

Auditory saltation is an illusion in which a train of clicks, the first half of which is presented at one location and the other half of which is presented from a second location, is perceived as originating not only from the anchor points, but also from locations between them. That is, intermediate members of the series of clicks have their spatial locations systematically misperceived. In the present study, auditory saltation was examined for the first time in the vertical midsagittal plane. Subjects rated the perceived continuity of motion for 8-click trains systematically varied in inter-click interval (ICI), direction of motion (up, down), and trial type ('saltation' versus 'real' motion). In all listeners, saltation stimuli supported robust saltation, but only for trials with ICIs less than about 120 ms. Real motion was rated as continuous for all ICIs. These data indicate that the auditory-saltation illusion can exploit monaural stimulus cues for source location in the generation of the illusory motion percept.