SUMMARY Pupil size is collectively controlled by the sympathetic dilator and parasympathetic sphincter muscles. Locus coeruleus (LC) activation has been shown to evoke pupil dilation, yet how the sympathetic and parasympathetic pathways contribute to this dilation remains unknown. We examined pupil dilation elicited by LC activation in lightly anesthetized rats. Unilateral LC activation evoked bilateral, yet lateralized pupil dilation, i.e, the ipsilateral dilation was significantly larger than the contralateral dilation. Surgically blocking the ipsilateral, but not contralateral, sympathetic pathway significantly reduced lateralization, suggesting lateralization is mainly due to sympathetic contribution. Moreover, we found that sympathetic, but not parasympathetic, contribution is correlated with LC activation frequency. Together, our results unveiled the frequency-dependent contributions of the sympathetic and parasympathetic pathways to LC-activation-evoked pupil dilation, and suggest that lateralization in task-evoked pupil dilations may be used as a biomarker for autonomic tone.

We investigated locus coeruleus (LC) modulation of thalamic feature selectivity through reverse correlation analysis of single-unit recordings from different stages of the rat vibrissa pathway. LC activation increased feature selectivity, drastically improving thalamic information transmission. We found this improvement was dependent on both local activation of α-adrenergic receptors and modulation of T-type calcium channels in the thalamus and was not due to LC modulation of trigeminothalamic feedforward or corticothalamic feedback inputs. Tonic spikes with LC stimulation carried 3-times the information than did tonic spikes without LC stimulation. Modelling confirmed norepinephrine (NE) regulation of intrathalamic circuit dynamics led to the improved information transmission. Behavioral data demonstrated that LC activation increased the perceptual performance of animals performing tactile discrimination tasks through LC-NE optimization of thalamic sensory processing. These results suggest a new sub-dimension within the tonic mode in which brain state can optimize thalamic sensory processing through modulation of intrathalamic circuit dynamics.

Oncolytic viruses pose many questions in their use in cancer therapy. In this study, we assessed the potential of mpJX-594 (mouse-prototype JX-594), a replication-competent vaccinia virus administered by intravenous injection, to target the tumor vasculature, produce immune activation and tumor cell killing more widespread than the infection, and suppress invasion and metastasis. These actions were examined in RIP-Tag2 transgenic mice with pancreatic neuroendocrine tumors that developed spontaneously and progressed as in humans. mpJX-594 initially infected tumor vascular endothelial cells, leading to vascular pruning and prolonged leakage in tumors but not in normal organs; parallel effects were observed in U87 gliomas. Viral infection spread to tumor cells, where tumor cell killing was much more widespread than the infection. Widespread tumor cell killing at 5 days was prevented by depletion of CD8 T lymphocytes and did not require GM-CSF, as mpJX-594 variants that expressed human, mouse, or no GM-CSF produced equivalent amounts of killing. The antivascular, antitumor, and antimetastatic effects of mpJX-594 were amplified by concurrent or sequential administration of sunitinib, a multitargeted receptor tyrosine kinase inhibitor. These effects were not mimicked by selective inhibition of VEGFR2 despite equivalent vascular pruning, but were accompanied by suppression of regulatory T cells and greater influx of activated CD8 T cells. Together, our results showed that mpJX-594 targets tumor blood vessels, spreads secondarily to tumor cells, and produces widespread CD8 T-cell-dependent tumor cell killing in primary tumors and metastases, and that these effects can be amplified by coadministration of sunitinib. These findings reveal multiple unrecognized features of the antitumor properties of oncolytic vaccinia viruses, all of which can be amplified by the multitargeted kinase inhibitor sunitinib. .

Non-luminance-mediated changes in pupil size have been widely used to index arousal state. Recent animal studies have demonstrated correlations between behavioral state-related pupil dynamics and sensory processing. However, the relationship between pupil-linked arousal and behavior in animals performing perceptual tasks has not been fully elucidated. In the present study, we trained head-fixed rats to discriminate between directions of whisker movements using a Go/No-Go discrimination paradigm while imaging their pupils. Reaction times in this discrimination task were significantly slower than in previously reported detection tasks with similar setup, suggesting that discrimination required an increased cognitive load. We found the pupils dilated for all trials following stimulus presentation. Interestingly, in correct rejection trials, where pupil dilations solely resulted from cognitive processing, dilations were larger for more difficult stimuli. Baseline pupil size before stimulus presentation strongly correlated with behavior, as perceptual sensitivity peaked at intermediate pupil baselines and reaction time was fastest at large baselines. We further explored these relationships by investigating to what extent pupil baseline was predictive of upcoming behavior and found that a Bayesian decoder had significantly greater-than-chance probability in correctly predicting behavioral outcomes. Moreover, the outcome of the previous trial showed a strong correlation with behavior on present trials. Animals were more liberal and faster in responding following hit trials, whereas perceptual sensitivity was greatest following correct rejection trials. Taken together, these results suggest a tight correlation between pupil dynamics, perceptual performance, and reaction time in behaving rats, all of which are modulated by fluctuating arousal state. NEW & NOTEWORTHY In this study, we for the first time demonstrated that head-fixed rats were able to discriminate different directions of whisker movement. Interestingly, we found that the pupil dilated more when discriminating more difficult stimuli, a phenomenon reported in human subjects but not in animals. Baseline pupil size before stimulus presentation was found to strongly correlate with behavior, and a Bayesian decoder had significantly greater-than-chance probability in correctly predicting behavioral outcomes based on the baseline pupil size.

Inhibition of vascular endothelial growth factor (VEGF) signaling can promote lymph node metastasis in preclinical models, but the mechanism is not fully understood, and successful methods of prevention have not been found. Signaling of hepatocyte growth factor (HGF) and its receptor c-Met can promote the growth of lymphatics and metastasis of some tumors. We sought to explore the contributions of c-Met signaling to lymph node metastasis after inhibition of VEGF signaling. In particular, we examined whether c-Met is upregulated in lymphatics in or near pancreatic neuroendocrine tumors in RIP-Tag2 transgenic mice and whether lymph node metastasis can be reduced by concurrent inhibition of VEGF and c-Met signaling. Inhibition of VEGF signaling by anti-VEGF antibody or sunitinib in mice from age 14 to 17 weeks was accompanied by more intratumoral lymphatics, more tumor cells inside lymphatics, and more lymph node metastases. Under these conditions, lymphatic endothelial cells - like tumor cells - had strong immunoreactivity for c-Met and phospho-c-Met. c-Met blockade by the selective inhibitor PF-04217903 significantly reduced metastasis to local lymph nodes. Together, these results indicate that inhibition of VEGF signaling in RIP-Tag2 mice upregulates c-Met expression in lymphatic endothelial cells, increases the number of intratumoral lymphatics and number of tumor cells within lymphatics, and promotes metastasis to local lymph nodes. Prevention of lymph node metastasis by PF-04217903 in this setting implicates c-Met signaling in tumor cell spread to lymph nodes.

In decision‐making tasks, neural circuits involved in different aspects of information processing may activate the central arousal system, likely through their interconnection with brainstem arousal nuclei, collectively contributing to the observed pupil‐linked phasic arousal. However, the individual components of the phasic arousal associated with different elements of information processing and their effects on behavior remain little known. In this study, we used machine learning techniques to decompose pupil‐linked phasic arousal evoked by different components of information processing in rats performing a Go/No‐Go perceptual decision‐making task. We found that phasic arousal evoked by stimulus encoding was larger for the Go stimulus than the No‐Go stimulus. For each session, the separation between distributions of phasic arousal evoked by the Go and by the No‐Go stimulus was predictive of perceptual performance. The separation between distributions of decision‐formation‐evoked arousal on correct and incorrect trials was correlated with decision criterion but not perceptual performance. When a Go stimulus was presented, the action of go was primarily determined by the phasic arousal evoked by stimulus encoding. On the contrary, when a No‐Go stimulus was presented, the action of go was determined by phasic arousal elicited by both stimulus encoding and decision formation. Drift diffusion modeling revealed that the four model parameters were better accounted for when phasic arousal elicited by both stimulus encoding and decision formation was considered. These results suggest that the interplay between phasic arousal evoked by both stimulus encoding and decision formation has important functional consequences on forming behavioral choice in perceptual decision‐making.

Several physiology signals, including heart rate and pupil size, have been widely used as peripheral indices of arousal to evaluate the effects of arousal on brain functions. However, whether behavior depends differently on arousal indexed by these physiological signals remains unclear. We simultaneously recorded electrocardiogram (ECG) and pupil size in head-fixed rats performing tactile discrimination tasks. We found both heartbeat dynamics and pupil size co-varied with behavioral outcomes, indicating behavior was dependent upon arousal indexed by the two physiological signals. To estimate the potential difference between the effects of pupil-linked arousal and heart rate-linked arousal on behavior, we constructed a Bayesian decoder to predict animals' behavior from pupil size and heart rate prior to stimulus presentation. The performance of the decoder was significantly better when using both heart rate and pupil size as inputs than when using either of them alone, suggesting the effects of the two arousal systems on behavior are not completely redundant. Supporting this notion, we found that, on a substantial portion of trials correctly predicted by the heart rate-based decoder, the pupil size-based decoder failed to correctly predict animals' behavior. Taken together, these results suggest that pupil-linked and heart rate-linked arousal systems exert different influences on animals' behavior.

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