Peptidergic and functional delineation of the Edinger-Westphal nucleus.

Many neuronal populations that release fast-acting excitatory and inhibitory neurotransmitters in the brain also contain slower-acting neuropeptides. These facultative peptidergic cell types are common, but it remains uncertain whether neurons that solely release peptides exist. Our fluorescence in situ hybridization, genetically targeted electron microscopy, and electrophysiological characterization suggest that most neurons of the non-cholinergic, centrally projecting Edinger-Westphal nucleus in mice are obligately peptidergic. We further show, using anterograde projection mapping, monosynaptic retrograde tracing, angled-tip fiber photometry, and chemogenetic modulation and genetically targeted ablation in conjunction with canonical assays for anxiety, that this peptidergic population activates in response to loss of motor control and promotes anxiety responses. Together, these findings elucidate an integrative, ethologically relevant role for the Edinger-Westphal nucleus and functionally align the nucleus with the periaqueductal gray, where it resides. This work advances our understanding of peptidergic modulation of anxiety and provides a framework for future investigations of peptidergic systems.

Downregulation of PACAP and the PAC1 Receptor in the Basal Ganglia, Substantia Nigra and Centrally Projecting Edinger-Westphal Nucleus in the Rotenone model of Parkinson's Disease.

Numerous in vitro and in vivo models of Parkinson's disease (PD) demonstrate that pituitary adenylate cyclase-activating polypeptide (PACAP) conveys its strong neuroprotective actions mainly via its specific PAC1 receptor (PAC1R) in models of PD. We recently described the decrease in PAC1R protein content in the basal ganglia of macaques in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) model of PD that was partially reversed by levodopa therapy. In this work, we tested whether these observations occur also in the rotenone model of PD in the rat. The rotarod test revealed motor skill deterioration upon rotenone administration, which was reversed by benserazide/levodopa (B/L) treatment. The sucrose preference test suggested increased depression level while the open field test showed increased anxiety in rats rendered parkinsonian, regardless of the received B/L therapy. Reduced dopaminergic cell count in the substantia nigra pars compacta (SNpc) diminished the dopaminergic fiber density in the caudate-putamen (CPu) and decreased the peptidergic cell count in the centrally projecting Edinger-Westphal nucleus (EWcp), supporting the efficacy of rotenone treatment. RNAscope in situ hybridization revealed decreased PACAP mRNA (Adcyap1) and PAC1R mRNA (Adcyap1r1) expression in the CPu, globus pallidus, dopaminergic SNpc and peptidergic EWcp of rotenone-treated rats, but no remarkable downregulation occurred in the insular cortex. In the entopeduncular nucleus, only the Adcyap1r1 mRNA was downregulated in parkinsonian animals. B/L therapy attenuated the downregulation of Adcyap1 in the CPu only. Our current results further support the evolutionarily conserved role of the PACAP/PAC1R system in neuroprotection and its recruitment in the development/progression of neurodegenerative states such as PD.

Oxytocin Receptors in the Mouse Centrally-projecting Edinger-Westphal Nucleus and their Potential Functional Significance for Thermoregulation.

The centrally-projecting Edinger-Westphal nucleus (EWcp) has been shown to contribute to regulation of multiple functions, including responses to stress and fear, attention, food consumption, addiction, body temperature and maternal behaviors. However, receptors involved in regulation of these behaviors through EWcp remain poorly characterized. On the other hand, the oxytocin peptide (OXT) is also known to regulate a substantial number of physiological responses and behaviors. Here we show that mRNA encoding OXT receptors (Oxtr) is expressed in EWcp of male and female C57BL/6J mice. These receptors are present on urocortin 1 (Ucn) mRNA-containing neurons and, to a lesser extent, on neurons in EWcp expressing the vesicular glutamate transporter 2 (Vglut2) mRNA of EWcp. Using RNAscope in situ hybridization, we show that neurons containing Ucn and Vglut2 mRNAs are two intermingled, but independent subpopulations in EWcp and characterize their relationship with other populations of neurons in the vicinity of this nucleus. Using immunohistochemistry, we show that intraperitoneal (IP) administration of OXT can induce FOS in Oxtr-containing neurons, suggesting that these receptors on EWcp neurons are functional. A follow up study showed that injection of OXT (2.3 or 7.7 mg/kg, IP) is accompanied by a decrease in body temperature. Since EWcp is known to be involved in regulation of body temperature, we hypothesize that OXT's effects on body temperature could be mediated through the EWcp. The contribution of OXTR in EWcp to regulation of various functions of EWcp and OXT needs to be deciphered.

Peptidergic neurons of the Edinger-Westphal nucleus express TRPA1 ion channel that is downregulated both upon chronic variable mild stress in male mice and in humans who died by suicide.

BACKGROUND: Transient receptor potential ankyrin 1 (TRPA1), a cation channel, is expressed predominantly in primary sensory neurons, but its central distribution and role in mood control are not well understood. We investigated whether TRPA1 is expressed in the urocortin 1 (UCN1)-immunoreactive centrally projecting Edinger-Westphal nucleus (EWcp), and we hypothesized that chronic variable mild stress (CVMS) would reduce its expression in mice. We anticipated that TRPA1 mRNA would be present in the human EWcp, and that it would be downregulated in people who died by suicide. METHODS: We exposed Trpa1 knockout and wild-type mice to CVMS or no-stress control conditions. We then performed behavioural tests for depression and anxiety, and we evaluated physical and endocrinological parameters of stress. We assessed EWcp Trpa1 and Ucn1 mRNA expression, as well as UCN1 peptide content, using RNA-scope in situ hybridization and immunofluorescence. We tested human EWcp samples for TRPA1 using reverse transcription polymerase chain reaction. RESULTS: Trpa1 mRNA was colocalized with EWcp/UCN1 neurons. Non-stressed Trpa1 knockout mice expressed higher levels of Ucn1 mRNA, had less body weight gain and showed greater immobility in the forced swim test than wild-type mice. CVMS downregulated EWcp/Trpa1 expression and increased immobility in the forced swim test only in wild-type mice. We confirmed that TRPA1 mRNA expression was downregulated in the human EWcp in people who died by suicide. LIMITATIONS: Developmental compensations and the global lack of TRPA1 may have influenced our findings. Because experimental data came from male brains only, we have no evidence for whether findings would be similar in female brains. Because a TRPA1-specific antibody is lacking, we have provided mRNA data only. Limited access to high-quality human tissues restricted sample size. CONCLUSION: TRPA1 in EWcp/UCN1 neurons might contribute to the regulation of depression-like behaviour and stress adaptation response in mice. In humans, TRPA1 might contribute to mood control via EWcp/UCN1 neurons.

Neurodegeneration in the centrally-projecting Edinger-Westphal nucleus contributes to the non-motor symptoms of Parkinson's disease in the rat.

BACKGROUND: The neuropathological background of major depression and anxiety as non-motor symptoms of Parkinson's disease is much less understood than classical motor symptoms. Although, neurodegeneration of the Edinger-Westphal nucleus in human Parkinson's disease is a known phenomenon, its possible significance in mood status has never been elucidated. In this work we aimed at investigating whether neuron loss and alpha-synuclein accumulation in the urocortin 1 containing (UCN1) cells of the centrally-projecting Edinger-Westphal (EWcp) nucleus is associated with anxiety and depression-like state in the rat. METHODS: Systemic chronic rotenone administration as well as targeted leptin-saporin-induced lesions of EWcp/UCN1 neurons were conducted. Rotarod, open field and sucrose preference tests were performed to assess motor performance and mood status. Multiple immunofluorescence combined with RNAscope were used to reveal the functional-morphological changes. Two-sample Student's t test, Spearman's rank correlation analysis and Mann-Whitney U tests were used for statistics. RESULTS: In the rotenone model, besides motor deficit, an anxious and depression-like phenotype was detected. Well-comparable neuron loss, cytoplasmic alpha-synuclein accumulation as well as astro- and microglial activation were observed both in the substantia nigra pars compacta and EWcp. Occasionally, UCN1-immunoreactive neuronal debris was observed in phagocytotic microglia. UCN1 peptide content of viable EWcp cells correlated with dopaminergic substantia nigra cell count. Importantly, other mood status-related dopaminergic (ventral tegmental area), serotonergic (dorsal and median raphe) and noradrenergic (locus ceruleus and A5 area) brainstem centers did not show remarkable morphological changes. Targeted partial selective EWcp/UCN1 neuron ablation induced similar mood status without motor symptoms. CONCLUSIONS: Our findings collectively suggest that neurodegeneration of urocortinergic EWcp contributes to the mood-related non-motor symptoms in toxic models of Parkinson's disease in the rat.

A Novel Tectal/Pretectal Population of Premotor Lens Accommodation Neurons.

Purpose: Under real-world conditions, saccades are often accompanied by changes in vergence angle and lens accommodation that compensate for changes in the distance between the current fixation point and the next target. As the superior colliculus directs saccades, we examined whether it contains premotor neurons that might control lens compensation for target distance. Methods: Rabies virus or recombinant rabies virus was injected into the ciliary bodies of Macaca fascicularis monkeys to label circuits controlling lens accommodation via retrograde transsynaptic transport. In addition, conventional anterograde tracers were used to confirm the rabies findings with respect to projections to preganglionic Edinger-Westphal motoneurons. Results: At time courses that rabies virus labeled lens-related premotor neurons in the supraoculomotor area and central mesencephalic reticular formation, labeled neurons were not found within the superior colliculus. They were, however, found bilaterally in the medial pretectal nucleus continuing caudally into the tectal longitudinal column, which lies on the midline, between the colliculi. A bilateral projection by this area to the preganglionic Edinger-Westphal nucleus was confirmed by anterograde tracing. Only at longer time courses were cells labeled in the superior colliculus. Conclusions: The superior colliculus does not provide premotor input to preganglionic Edinger-Westphal nucleus motoneurons, but may provide input to lens-related premotor populations in the supraoculomotor area and central mesencephalic reticular formation. There is, however, a novel third population of lens-related premotor neurons in the tectal longitudinal column and rostrally adjacent medial pretectal nucleus. The specific function of this premotor population remains to be determined.

Leptin coordinates efferent sympathetic outflow to the white adipose tissue through the midbrain centrally-projecting Edinger-Westphal nucleus in male rats.

The centrally-projecting Edinger-Westphal nucleus (EWcp) hosts a large population of neurons expressing urocortin 1 (Ucn1) and about half of these neurons also express the leptin receptor (LepRb). Previously, we have shown that the peripheral adiposity hormone leptin signaling energy surfeit modulates EWcp neurons' activity. Here, we hypothesized that Ucn1/LepRb neurons in the EWcp would act as a crucial neuronal node in the brain-white adipose tissue (WAT) axis modulating efferent sympathetic outflow to the WAT. We showed that leptin bound to neurons of the EWcp stimulated STAT3 phosphorylation, and increased Ucn1-production in a time-dependent manner. Besides, retrograde transneuronal tract-tracing using pseudorabies virus (PRV) identified EWcp Ucn1 neurons connected to WAT. Interestingly, reducing EWcp Ucn1 contents by ablating EWcp LepRb-positive neurons with leptin-saporin, did not affect food intake and body weight gain, but substantially (+26%) increased WAT weight accompanied by a higher plasma leptin level and changed plasma lipid profile. We also found that ablation of EWcp Ucn1/LepRb neurons resulted in lower respiratory quotient and oxygen consumption one week after surgery, but was comparable to sham values after 3 and 5 weeks of surgery. Taken together, we report that EWcp/LepRb/Ucn1 neurons not only respond to leptin signaling but also control WAT size and fat metabolism without altering food intake. These data suggest the existence of a EWcp-WAT circuitry allowing an organism to recruit fuels without being able to eat in situations such as the fight-or-flight response.

Vesicular glutamate transporter 2-containing neurons of the centrally-projecting Edinger-Westphal nucleus regulate alcohol drinking and body temperature.

Previous studies in rodents have repeatedly demonstrated that the centrally-projecting Edinger-Westphal nucleus (EWcp) is highly sensitive to alcohol and is also involved in regulating alcohol intake and body temperature. Historically, the EWcp has been known as the main site of Urocortin 1 (Ucn1) expression, a corticotropin-releasing factor-related peptide, in the brain. However, the EWcp also contains other populations of neurons, including neurons that express the vesicular glutamate transporter 2 (Vglut2). Here we transduced the EWcp with adeno-associated viruses (AAVs) encoding Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) to test the role of the EWcp in alcohol drinking and in the regulation of body temperature. Activation of the EWcp with excitatory DREADDs inhibited alcohol intake in a 2-bottle choice procedure in male C57BL/6J mice, whereas inhibition of the EWcp with DREADDs had no effect. Surprisingly, analysis of DREADD expression indicated Ucn1-containing neurons of the EWcp did not express DREADDs. In contrast, AAVs transduced non-Ucn1-containing EWcp neurons. Subsequent experiments showed that the inhibitory effect of EWcp activation on alcohol intake was also present in male Ucn1 KO mice, suggesting that a Ucn1-devoid population of EWcp regulates alcohol intake. A final set of chemogenetic experiments showed that activation of Vglut2-expressing EWcp neurons inhibited alcohol intake and induced hypothermia in male and female mice. These studies expand on previous literature by indicating that a glutamatergic, Ucn1-devoid subpopulation of the EWcp regulates alcohol consumption and body temperature.

Magnocellular and Parvocellular Mediated Luminance Contrast Discrimination in Severe Alcohol Use Disorder.

BACKGROUND: Severe alcohol use disorder (SAUD) is associated with widespread cognitive impairments, including low-level visual processing deficits that persist after prolonged abstinence. However, the extent and characteristics of these visual deficits remain largely undetermined, impeding the identification of their underlying mechanisms and influence on higher-order processing. In particular, little work has been conducted to assess the integrity of the magnocellular (MC) and parvocellular (PC) visual pathways, namely the 2 main visual streams that convey information from the retina up to striate, extrastriate, and dorsal/ventral cerebral regions. METHODS: We investigated achromatic luminance contrast processing mediated by inferred MC and PC pathways in 33 patients with SAUD and 32 matched healthy controls using 2 psychophysical pedestal contrast discrimination tasks that promote responses of inferred MC or PC pathways. We relied on a staircase procedure to assess participants' ability to detect small changes in luminance within an array of 4 gray squares that were either continuously presented (steady pedestal, MC-biased) or briefly flashed (pulsed pedestal, PC-biased). RESULTS: We replicated the expected pattern of MC and PC contrast responses in healthy controls. We found preserved dissociation of MC and PC contrast signatures in SAUD but higher MC-mediated mean contrast discrimination thresholds combined with a steeper PC-mediated contrast discrimination slope compared with healthy controls. CONCLUSION: These findings indicate altered MC-mediated contrast sensitivity and PC-mediated contrast gain, confirming the presence of early sensory disturbances in individuals with SAUD. Such low-level deficits, while usually overlooked, might influence higher-order abilities (e.g., memory, executive functions) in SAUD by disturbing the "coarse-to-fine" tuning of the visual system, which relies on the distinct functional properties of MC and PC pathways and ensures proper and efficient monitoring of the environment.

Is Primate Lens Accommodation Unilaterally or Bilaterally Controlled?

Purpose: In frontal-eyed mammals such as primates, eye movements are coordinated so that the lines of sight are directed at targets in a manner that adjusts for target distance. The lens of each eye must also be adjusted with respect to target distance to maintain precise focus. Whether the systems for controlling eye movements are monocularly or binocularly organized is currently a point of contention. We recently determined that the premotor neurons controlling the lens of one eye are bilaterally distributed in the midbrain. In this study, we examine whether this is due to premotor neurons projecting bilaterally to the preganglionic Edinger-Westphal nuclei, or by a mixture of ipsilaterally and contralaterally projecting cells supplying each nucleus. Methods: The ciliary muscles of Macaca fasicularis monkeys were injected with recombinant forms of the N2c rabies virus, one eye with virus that produced a green fluorescent marker and the other eye with a virus that produced a red fluorescent marker. Results: Preganglionic motoneurons in the Edinger-Westphal nucleus displayed the same marker as the ipsilateral injected muscle. Many of the premotor neurons in the supraoculomotor area and central mesencephalic reticular formation were doubly labeled. Others were labeled from either the ipsilateral or contralateral eye. Conclusions: These results suggest that both monocular control and binocular control of lens accommodation are present. Binocular inputs yoke the accommodation in the two eyes. Monocular inputs may allow modification related to differences in each eye's target distance or differences in the capacities of the two ciliary muscles.

Spatial frequency tuning of motor responses reveals differential contribution of dorsal and ventral systems to action comprehension.

Understanding object-directed actions performed by others is central to everyday life. This ability is thought to rely on the interaction between the dorsal action observation network (AON) and a ventral object recognition pathway. On this view, the AON would encode action kinematics, and the ventral pathway, the most likely intention afforded by the objects. However, experimental evidence supporting this model is still scarce. Here, we aimed to disentangle the contribution of dorsal vs. ventral pathways to action comprehension by exploiting their differential tuning to low-spatial frequencies (LSFs) and high-spatial frequencies (HSFs). We filtered naturalistic action images to contain only LSF or HSF and measured behavioral performance and corticospinal excitability (CSE) using transcranial magnetic stimulation (TMS). Actions were embedded in congruent or incongruent scenarios as defined by the compatibility between grips and intentions afforded by the contextual objects. Behaviorally, participants were better at discriminating congruent actions in intact than LSF images. This effect was reversed for incongruent actions, with better performance for LSF than intact and HSF. These modulations were mirrored at the neurophysiological level, with greater CSE facilitation for congruent than incongruent actions for HSF and the opposite pattern for LSF images. Finally, only for LSF did we observe CSE modulations according to grip kinematics. While results point to differential dorsal (LSF) and ventral (HSF) contributions to action comprehension for grip and context encoding, respectively, the negative congruency effect for LSF images suggests that object processing may influence action perception not only through ventral-to-dorsal connections, but also through a dorsal-to-dorsal route involved in predictive processing.

Temporal information loss in the macaque early visual system.

Stimuli that modulate neuronal activity are not always detectable, indicating a loss of information between the modulated neurons and perception. To identify where in the macaque visual system information about periodic light modulations is lost, signal-to-noise ratios were compared across simulated cone photoreceptors, lateral geniculate nucleus (LGN) neurons, and perceptual judgements. Stimuli were drifting, threshold-contrast Gabor patterns on a photopic background. The sensitivity of LGN neurons, extrapolated to populations, was similar to the monkeys' at low temporal frequencies. At high temporal frequencies, LGN sensitivity exceeded the monkeys' and approached the upper bound set by cone photocurrents. These results confirm a loss of high-frequency information downstream of the LGN. However, this loss accounted for only about 5% of the total. Phototransduction accounted for essentially all of the rest. Together, these results show that low temporal frequency information is lost primarily between the cones and the LGN, whereas high-frequency information is lost primarily within the cones, with a small additional loss downstream of the LGN.

Effects of pharmacological inhibition of the centrally-projecting Edinger-Westphal nucleus on ethanol-induced conditioned place preference and body temperature.

Alcohol use disorder is a chronic disease characterized in part by repeated relapsing events. Exposure to environmental stimuli or cues that have previously been associated with the effects of alcohol can promote relapse through the triggering of craving for alcohol. Therefore, identifying and characterizing neuronal populations that may regulate these associations is of the upmost importance. Previous studies have implicated the centrally-projecting Edinger Westphal nucleus (EWcp) in this process, as the EWcp is both sensitive to, and can regulate alcohol intake. To date however, it is unclear if the EWcp is involved in the formation or expression of these alcohol-cue associations. As such, the present studies examined the involvement of the EWcp in male DBA/2J mice in the acquisition and expression of place preference for an alcohol-paired cue using the conditioned place preference (CPP) procedure. Pharmacological inhibition of the EWcp via the GABAA and GABAB receptor agonists muscimol and baclofen did not affect either the acquisition or the expression of CPP. Follow up studies did find however, that pharmacological inhibition of the EWcp increased body temperature and prevented alcohol-induced increases in c-Fos expression in the EWcp. When considered in light of previous studies, the present results indicate that the EWcp may be involved in the regulation of alcohol self-administration, and not conditioned alcohol-seeking. Additionally, the present studies provide further evidence for the involvement of the EWcp in thermoregulation and help elucidate the molecular mechanisms by which alcohol increases c-Fos in the EWcp.

Pupillary light reflex circuits in the Macaque Monkey: the olivary pretectal nucleus.

The olivary pretectal nucleus is the first central connection in the pupillary light reflex pathway, the circuit that adjusts the diameter of the pupil in response to ambient light levels. This study investigated aspects of the morphology and connectivity of the olivary pretectal nucleus in macaque monkeys by use of anterograde and retrograde tracers. Within the pretectum, the vast majority of neurons projecting to the preganglionic Edinger-Westphal nucleus were found within the olivary pretectal nucleus. Most of these neurons had somata located at the periphery of the nucleus and their heavily branched dendrites extended into the core of the nucleus. Retinal terminals were concentrated within the borders of the olivary pretectal nucleus. Ultrastructural examination of these terminals showed that they had clear spherical vesicles, occasional dense-core vesicles, and made asymmetric synaptic contacts. Retrogradely labeled cells projecting to the preganglionic Edinger-Westphal nucleus displayed relatively few somatic contacts. Double labeling indicated that these neurons receive direct retinal input. The concentration of retinal terminals within the nucleus and the extensive dendritic trees of the olivary projection cells provide a substrate for very large receptive fields. In some species, pretectal commissural connections are a substrate for balancing the direct and consensual pupillary responses to produce pupils of equal size. In the macaque, there was little evidence for such a commissural projection based on either anterograde or retrograde tracing. This may be due to the fact that each macaque retina provides nearly equal density projections to the ipsilateral and contralateral olivary pretectal nucleus.

Pupillary light reflex circuits in the macaque monkey: the preganglionic Edinger-Westphal nucleus.

The motor outflow for the pupillary light reflex originates in the preganglionic motoneuron subdivision of the Edinger-Westphal nucleus (EWpg), which also mediates lens accommodation. Despite their importance for vision, the morphology, ultrastructure and luminance-related inputs of these motoneurons have not been fully described in primates. In macaque monkeys, we labeled EWpg motoneurons from ciliary ganglion and orbital injections. Both approaches indicated preganglionic motoneurons occupy an EWpg organized as a unitary, ipsilateral cell column. When tracers were placed in the pretectal complex, labeled terminals targeted the ipsilateral EWpg and reached contralateral EWpg by crossing both above and below the cerebral aqueduct. They also terminated in the lateral visceral column, a ventrolateral periaqueductal gray region containing neurons projecting to the contralateral pretectum. Combining olivary pretectal and ciliary ganglion injections to determine whether a direct pupillary light reflex projection is present revealed a labeled motoneuron subpopulation that displayed close associations with labeled pretectal terminal boutons. Ultrastructurally, this subpopulation received synaptic contacts from labeled pretectal terminals that contained numerous clear spherical vesicles, suggesting excitation, and scattered dense-core vesicles, suggesting peptidergic co-transmitters. A variety of axon terminal classes, some of which may serve the near response, synapsed on preganglionic motoneurons. Quantitative analysis indicated that pupillary motoneurons receive more inhibitory inputs than lens motoneurons. To summarize, the pupillary light reflex circuit utilizes a monosynaptic, excitatory, bilateral pretectal projection to a distinct subpopulation of EWpg motoneurons. Furthermore, the interconnections between the lateral visceral column and olivary pretectal nucleus may provide pretectal cells with bilateral retinal fields.

Fos activation patterns related to acute ethanol and conditioned taste aversion in adolescent and adult rats.

Studies in rats have revealed marked age differences in sensitivity to the aversive properties of ethanol, with a developmental insensitivity to ethanol aversion that is most pronounced during pre- and early adolescence, declining thereafter to reach the enhanced aversive sensitivity of adults. The adolescent brain undergoes significant transitions throughout adolescence, including in regions linked with drug reward and aversion; however, it is unknown how ontogenetic changes within this reward/aversion circuitry contribute to developmental differences in aversive sensitivity. The current study examined early adolescent (postnatal day [P]28-30) and adult (P72-74) Sprague-Dawley male rats for conditioned taste aversion (CTA) after doses of 0, 1.0, or 2.5 g/kg ethanol, and patterns of neuronal activation in response to ethanol using Fos-like immunohistochemistry (Fos+) to uncover regions where age differences in activation are associated with ethanol aversion. An adolescent-specific ethanol-induced increase in Fos+ staining was seen within the nucleus accumbens shell and core. An age difference was also noted within the Edinger-Westphal nucleus (EW) following administration of the lower dose of ethanol, with 1 g/kg ethanol producing CTA in adults but not in adolescents and inducing a greater EW Fos response in adults than adolescents. Regression analysis revealed that greater numbers of Fos+ neurons within the EW and insula (Ins) were related to lower consumption of the conditioned stimulus (CS) on test day (reflecting greater CTA). Some regionally specific age differences in Fos+ were noted under baseline conditions, with adolescents displaying fewer Fos+ neurons than adults within the prelimbic (PrL) cortex, but more than adults in the bed nucleus of the stria terminalis (BNST). In the BNST (but not PrL), ethanol-induced increases in Fos-immunoreactivity (IR) were evident at both ages. Increased ethanol-induced activity within critical appetitive brain regions (NAc core and shell) supports a role for greater reward-related activation during adolescence, possibly along with attenuated responsiveness to ethanol in EW and Ins in the age-typical resistance of adolescents to the aversive properties of ethanol.

Serotonin receptor 2c-expressing cells in the ventral CA1 control attention via innervation of the Edinger-Westphal nucleus.

The hippocampus is divided into dorsal and ventral zones along its principal axis. The dorsal hippocampus is critical for learning and memory, yet the basic function of the ventral hippocampus remains elusive. Here we genetically manipulate a subset of excitatory neurons expressing the serotonin receptor 2c (Htr2c) in the ventral hippocampus. Genetically modified virus tracing reveals that these Htr2c cells establish monosynaptic excitatory connections with newly identified neurons in the Edinger-Westphal nucleus (EW), which directly innervate the medial prefrontal cortex. Inactivation of Htr2c cells impairs behavioral performance in a visual-detection task that demands attention, without affecting novel-object recognition, learning, or memory. This attention deficit was recapitulated by inhibition of EW cells and rescued by activation of EW cells or synaptic projections from Htr2c cells onto EW cells. This study uncovers a synaptic pathway for control of attention.

The effects of red surrounds on visual magnocellular and parvocellular cortical processing and perception.

More than 50 years ago, Hubel and Wiesel identified a subpopulation of geniculate magnocellular (M) neurons that are suppressed by diffuse red light. Since then, many human psychophysical studies have used red and green backgrounds to study the effects of M suppression on visual task performance, as a means to better understand neurodevelopmental disorders such as dyslexia and schizophrenia. Few of these studies have explicitly assessed the relative effects of red backgrounds on the M and P (parvocellular) pathways. Here we compared the effects of red and green diffuse background illumination on well-accepted cortical M and P signatures, both physiologically through nonlinear analysis of visual evoked potentials (VEPs; N = 15), and psychophysically through pulsed and steady pedestal perceptual thresholds (N = 9 with gray pedestals and N = 8 with colored pedestals). Red surrounds reduced P-generated temporal nonlinearity in the VEPs, but they did not influence M-generated VEP signatures. The steady and pulsed pedestal results suggest that red surrounds can have different effects on M and P contrast sensitivities, depending on whether the target is colored gray or red, presented centrally or peripherally, or whether it is brighter or dimmer than the surround. Our results highlight difficulties in interpreting the effects of red backgrounds on human VEPs or perception in terms of M specific suppression.

Activity-based anorexia activates CRF immunoreactive neurons in female rats.

Activity-based anorexia (ABA) is a well-established animal model mimicking the eating disorder anorexia nervosa (AN). Since the pathophysiology of AN is yet poorly understood and specific drug treatments are lacking so far, animal models might be useful to further understand this disease. ABA consists of time-restricted access to food for 1.5 h/day and the possibility to exercise in a running wheel for 24 h/day. This combination leads to robust body weight loss as observed in AN. Here, we investigated the activation of brain corticotropin-releasing factor (CRF) neurons, a transmitter involved in the response to stress, emotional processes and also food intake. After development of ABA, rat brains were processed for c-Fos and CRF double immunohistochemistry. ABA increased the number of c-Fos/CRF double labeled neurons in the paraventricular nucleus (PVN) and the dorsomedial hypothalamic nucleus (DMH) compared to the ad libitum (AL, ad libitum fed, no running wheel) and activity (AC, ad libitum fed, running wheel, p < 0.05) but not to the restricted feeding (RF, food for 1.5 h/day, no running wheel, p > 0.05) group. Also the number of CRF neurons was increased in the DMH of ABA rats compared to AL and AC (p < 0.05). In the Edinger-Westphal nucleus (EW) the number of c-Fos positive neurons was increased in ABA and RF compared to AC (p < 0.05), while the number of double labeled neurons was not different (p > 0.05). Taken together, brain CRF activated under conditions of ABA might play a role in the development and maintenance of this animal model and possibly also in human AN.

Desacyl Ghrelin Decreases Anxiety-like Behavior in Male Mice.

Ghrelin is a 28-amino acid polypeptide that regulates feeding, glucose metabolism, and emotionality (stress, anxiety, and depression). Plasma ghrelin circulates as desacyl ghrelin (DAG) or, in an acylated form, acyl ghrelin (AG), through the actions of ghrelin O-acyltransferase (GOAT), exhibiting low or high affinity, respectively, for the growth hormone secretagogue receptor (GHSR) 1a. We investigated the role of endogenous AG, DAG, and GHSR1a signaling on anxiety and stress responses using ghrelin knockout (Ghr KO), GOAT KO, and Ghsr stop-floxed (Ghsr null) mice. Behavioral and hormonal responses were tested in the elevated plus maze and light/dark (LD) box. Mice lacking both AG and DAG (Ghr KO) increased anxiety-like behaviors across tests, whereas anxiety reactions were attenuated in DAG-treated Ghr KO mice and in mice lacking AG (GOAT KO). Notably, loss of GHSR1a (Ghsr null) did not affect anxiety-like behavior in any test. Administration of AG and DAG to Ghr KO mice with lifelong ghrelin deficiency reduced anxiety-like behavior and decreased phospho-extracellular signal-regulated kinase phosphorylation in the Edinger-Westphal nucleus in wild-type mice, a site normally expressing GHSR1a and involved in stress- and anxiety-related behavior. Collectively, our data demonstrate distinct roles for endogenous AG and DAG in regulation of anxiety responses and suggest that the behavioral impact of ghrelin may be context dependent.