Sleep dysregulation in binge eating disorder and "food addiction": the orexin (hypocretin) system as a potential neurobiological link.

It has been proposed that binge eating reflects a pathological compulsion driven by the "addictive" properties of foods. Proponents of this argument highlight the large degree of phenomenological and diagnostic overlap between binge eating disorder (BED) and substance use disorders (SUDs), including loss of control over how much is consumed and repeated unsuccessful attempts to abstain from consumption, as well as commonalities in brain structures involved in food and drug craving. To date, very little attention has been given to an additional behavioral symptom that BED shares with SUDs-sleep dysregulation-and the extent to which this may contribute to the pathophysiology of BED. Here, we review studies examining sleep outcomes in patients with BED, which collectively point to a heightened incidence of sleep abnormalities in BED. We identify the orexin (hypocretin) system as a potential neurobiological link between compulsive eating and sleep dysregulation in BED, and provide a comprehensive update on the evidence linking this system to these processes. Finally, drawing on evidence from the SUD literature indicating that the orexin system exhibits significant plasticity in response to drugs of abuse, we hypothesize that chronic palatable food consumption likewise increases orexin system activity, resulting in dysregulated sleep/wake patterns. Poor sleep, in turn, is predicted to exacerbate binge eating, contributing to a cycle of uncontrolled food consumption. By extension, we suggest that pharmacotherapies normalizing orexin signaling, which are currently being trialed for the treatment of SUDs, might also have utility in the clinical management of BED.

Orexin-1 Receptor Signaling in Ventral Pallidum Regulates Motivation for the Opioid Remifentanil.

Signaling at the orexin-1 receptor (OxR1) is important for motivated drug taking. Using a within-session behavioral economics (BE) procedure, we previously found that pharmacologic blockade of the OxR1 decreased motivation (increased demand elasticity) for the potent and short-acting opioid remifentanil and reduced low-effort remifentanil consumption. However, the mechanism through which orexin regulates remifentanil demand is currently unknown. Previous work implicated OxR1 signaling within ventral pallidum (VP) as a potential target. VP is densely innervated by orexin fibers and is known to regulate opioid reward. Accordingly, this study sought to determine the role of VP OxR1 signaling in remifentanil demand and cue-induced reinstatement of remifentanil seeking in male rats. Intra-VP microinjections of the OxR1 antagonist SB-334867 (SB) decreased motivation (increased demand elasticity; α) for remifentanil without affecting remifentanil consumption at low effort. Baseline α values predicted the degree of cue-induced remifentanil seeking, and microinjection of SB into VP attenuated this behavior without affecting extinction responding. Baseline α values also predicted SB efficacy, such that SB was most effective in attenuating reinstatement behavior in highly motivated rats. Together, these findings support a selective role for VP OxR1 signaling in motivation for the opioid remifentanil. Our findings also highlight the utility of BE in predicting relapse propensity and efficacy of treatment with OxR1 antagonists.SIGNIFICANCE STATEMENT Abuse of opioids has risen rapidly and continues to be a major health crisis. Thus, there is an urgent need to better understand the neurobiological and behavioral mechanisms underlying opioid addiction. Here, we investigate the role of orexin-1 receptor signaling (OxR1) within ventral pallidum (VP) in remifentanil demand and cue-induced reinstatement of remifentanil seeking. Using a within-session behavioral economics procedure, we show that intra-VP microinjections of the OxR1 antagonist SB-334867 decreased motivation (increased demand elasticity) without affecting remifentanil consumption at low effort. We also found that SB microinjected intra-VP attenuated cue-induced reinstatement of remifentanil seeking. Together, our results support a role for VP OxR1 signaling in opioid reward.

Increased Number and Activity of a Lateral Subpopulation of Hypothalamic Orexin/Hypocretin Neurons Underlies the Expression of an Addicted State in Rats.

BACKGROUND: The orexin (hypocretin) system is important for reward-driven motivation but has not been implicated in the expression of a multiphenotype addicted state. METHODS: Rats were assessed for economic demand for cocaine before and after 14 days of short access, long access, or intermittent access (IntA) to cocaine. Rats were also assessed for a number of other DSM-5-relevant addiction criteria following differential access conditions. Orexin system function was assessed by quantification of numbers and activity of orexin cells, pharmacological blockade of the orexin-1 receptor, and subregion-specific knockdown of orexin cell populations. RESULTS: IntA produced a cluster of addiction-like behaviors that closely recapitulate key diagnostic criteria for addiction to a greater extent than long access or short access. IntA was accompanied by an increase in number and activity of orexin-expressing neurons within the lateral hypothalamic subregion. This increase in orexin cell number and activity persisted during protracted withdrawal from cocaine for at least 150 days and was accompanied by enhanced incubation of craving in the same rats. Selective knockdown of lateral hypothalamic orexin neurons reduced motivation for cocaine, and orexin-1 receptor signaling played a larger role in drug seeking after IntA. CONCLUSIONS: We provide the first evidence that lateral hypothalamic orexin system function extends beyond general reward seeking to play a critical role in expression of a multiphenotype addiction-like state. Thus, the orexin system is a potential novel target for pharmacotherapies designed to treat cocaine addiction. In addition, these data point to the IntA model as a preferred approach to modeling addiction-like behavior in rats.

Demand elasticity predicts addiction endophenotypes and the therapeutic efficacy of an orexin/hypocretin-1 receptor antagonist in rats.

Behavioral economics is a powerful, translational approach for measuring drug demand in both humans and animals. Here, we asked if demand for cocaine in rats with limited drug experience could be used to identify individuals most at risk of expressing an addiction phenotype following either long- or intermittent access self-administration schedules, both of which model the transition to uncontrolled drug-seeking. Because the orexin-1 receptor antagonist SB-334867 (SB) is particularly effective at reducing drug-seeking in highly motivated individuals, we also asked whether demand measured after prolonged drug experience could predict SB efficacy. Demand elasticity (α) measured immediately following acquisition of cocaine self-administration ('baseline α') was positively correlated with α assessed after 2w of long- or intermittent access. Baseline α also predicted the magnitude of compulsive responding for cocaine, drug-seeking in initial abstinence and cued reinstatement following long-, intermittent- or standard short access. When demand was measured after these differential access conditions, α predicted the same addiction endophenotypes predicted by baseline α, as well as primed reinstatement and the emergence of negative emotional mood behavior following abstinence. α also predicted the efficacy of SB, such that high demand rats showed greater reductions in motivation for cocaine following SB compared to low demand rats. Together, these findings indicate that α might serve as a behavioral biomarker to predict individuals most likely to progress from controlled to uncontrolled drug use, and to identify individuals most likely to benefit from orexin-based therapies for the treatment of addiction.

Extracellular matrix nitration alters growth factor release and activates bioactive complement in human retinal pigment epithelial cells.

PURPOSE: We have shown previously that non-enzymatic nitration (NEN) of the extracellular matrix (ECM), which serves as a model of Bruch's membrane (BM) aging, has a profound effect on the behavior of the overlying retinal pigment epithelial (RPE) cells, including altered phagocytic ability, reduced cell adhesion, and inhibition of proliferation. We know that transplanted RPE monolayers will encounter a hostile sub-RPE environment, including age-related alterations in BM that may compromise cell function and survival. Here we use our previous NEN model of BM aging to determine the effects of NEN of the ECM on growth factor release and complement activation in RPE cells. METHODS: Human induced-pluripotent stem cells (iPSCs) were differentiated into RPE cells, and confirmed by immunohistochemistry, confocal microscopy, and polymerase chain reaction. IPSC-derived RPE cells were plated onto RPE-derived ECM under untreated or nitrite-modified conditions. Cells were cultured for 7 days and barrier function measured by transepithelial resistance (TER). Vascular endothelial growth factor (VEGF), pigment epithelium-derived factor (PEDF), and complement component C3a were measured using enzyme-linked immunosorbent assay (ELISA). RESULTS: On average nitrite-modified ECM increased VEGF release both apically and basally by 0.15 ± 0.014 ng/mL (p <0.0001) and 0.21 ± 0.022 ng/mL (p <0.0001), respectively, in iPSC-derived RPE cells. Nitrite-modified ECM increased PEDF release in iPSC-derived RPE cells apically by 0.16 ± 0.031 ng/mL (p <0.0001), but not basally (0.27 ± 0.015 vs. 0.32 ± 0.029 ng/mL, (p >0.05)). Nitrite-modified ECM increased production of C3a in iPSC-derived RPE cells by 0.52 ± 0.123 ng/mL (p <0.05). CONCLUSION: Nitrite-modified ECM increased VEGF, PEDF release, and C3a production in human iPSC-derived RPE cells. This model demonstrates changes seen in the basement membrane can lead to alterations in the cell biology of the RPE cells that may be related to the development of age-related macular degeneration.

New directions for the treatment of depression: Targeting the photic regulation of arousal and mood (PRAM) pathway.

Both preclinical and clinical studies demonstrate that depression is strongly associated with reduced light availability, which in turn contributes to decreased function of brain regions that control mood. Here, we review findings that support a critical pathway for the control of mood that depends upon ambient light. We put forward a novel hypothesis, functionally linking retina to locus coeruleus (LC) in depression, and discuss the role of norepinephrine in affective disease. Finally, we discuss how utilizing the chemogenetic tool Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) to precisely control this retina-LC circuit may be used as a novel therapeutic to treat depression.

Imaging mass spectrometry of the visual system: Advancing the molecular understanding of retina degenerations.

Visual sensation is fundamental for quality of life, and loss of vision to retinal degeneration is a debilitating condition. The eye is the only part of the central nervous system that can be noninvasively observed with optical imaging. In the clinics, various spectroscopic methods provide high spatial resolution images of the fundus and the developing degenerative lesions. However, the currently utilized tools are not specific enough to establish the molecular underpinnings of retinal diseases. In contrast, mass spectrometric imaging (MSI) is a powerful tool to identify molecularly specific disease indicators and classification markers. This technique is particularly well suited to the eye, where molecular information can be correlated with clinical data collected via noninvasive diagnostic imaging modalities. Recent studies during the last few recent years have uncovered a plethora of new spatially defined molecular information on several vision-threatening diseases, including age-related macular degeneration, Stargardt disease, glaucoma, cataract, as well as lipid disorders. Even though MS inside the eye cannot be performed noninvasively, by linking diagnostic and molecular information, these studies are the first step toward the development of smart ophthalmic diagnostic and surgical tools. Here, we provide an overview of current approaches applying MSI technology to ocular pathology.

Differentiation of Human Protein-Induced Pluripotent Stem Cells toward a Retinal Pigment Epithelial Cell Fate.

Compared with many induced pluripotent stem cell (iPSC) lines generated using retrovirus and other non-integrating methods, the utilization of human protein-induced iPSC (piPSC) lines may provide a safer alternative for the generation of retinal pigment epithelial (RPE) cells for transplantation in retinal degenerative diseases. Here we assess the ability of piPSCs to differentiate into RPE cells, and to perform native RPE cell behavior. piPSCs were seeded in 6-well low-attachment plates to allow embryoid body formation, and then analyzed for pluripotent stem cell markers NANOG, SSEA4 and TRA-1-60 by immunofluorescence. Following colony formation, piPSCs were assessed for confirmation of RPE cell differentiation by staining for zonula occludens (ZO-1), bestrophin, microphthalmia-associated transcription factor (MITF) and retinal pigment epithelium specific protein-65 (RPE65). To evaluate piPSC-RPE cell phagocytic ability, adult bovine photoreceptor rod outer segments (ROS) were fed to piPSC-RPE cells, which were analyzed by fluorescent microscopy and flow cytometry. Undifferentiated piPSCs expressed all pluripotent markers assessed and formed embryoid body aggregates after 7 days. Differentiated piPSC-RPE cells expressed ZO-1, bestrophin, MITF and RPE65, typical RPE cell markers. Flow cytometry revealed robust ingestion of fluorescently-labeled ROS by piPSC-RPE cells, which was over four-times greater than that of undifferentiated piPSCs and comparable to that of an immortalized RPE cell line. Phagocytosis activity by piPSC-RPE cells was significantly reduced after the addition of anti-integrin αVß5. In conclusion, piPSCs can be differentiated toward an RPE cell fate, expressing RPE cell markers and resembling native RPE cells in behavior. These results demonstrate that piPSCs can be differentiated into RPE-like cells using a method that has an increased safety profile, a critical consideration for the development of better treatments for retinal degenerative diseases such as age-related macular degeneration (AMD).

Retinoid Processing in Induced Pluripotent Stem Cell-Derived Retinal Pigment Epithelium Cultures.

Stem cell therapy for retinal degenerative diseases such as age-related macular degeneration is a promising clinical option for the replacement of photoreceptors and retinal pigment epithelium (RPE). Induced pluripotent stem cell technology has emerged as a viable potential source of cells for transplantation in retinal degenerative disorders. Induced pluripotent stem cells have been used to derive RPE and have been tested for their functional behavior. These cells have the ability to express RPE-specific proteins and morphologically resemble native RPE. Induced pluripotent stem cell-derived RPE are also able to contribute to the visual cycle by their ability to metabolize all-trans retinol, a critical function of RPE in maintaining visual function. Advances in induced pluripotent stem cell technology will contribute to the development of clinical therapies for retinal degenerative diseases as well as provide a tool to understand the pathology of these disorders.

Nitrite Modification of Extracellular Matrix Alters CD46 Expression and VEGF Release in Human Retinal Pigment Epithelium.

PURPOSE: Loss of CD46 has recently been implicated in choroidal neovascularization in mice. Herein we investigated the effect of nitrite modification of the extracellular matrix (ECM) as an in vitro model of "aging" and its effect on CD46 expression and vascular endothelial growth factor (VEGF) release in cocultured human retinal pigment epithelium (RPE). METHODS: ARPE-19 cells were plated onto RPE-derived ECM conditions (untreated; nitrite modified; nitrite modified followed by washing with Triton X-100; or nitrite modified followed by washing with Triton X-100 and coated with extracellular matrix ligands). Cells were cultured for 7 days and CD46 expression was analyzed by immunohistochemistry and Western blot. Additionally, CD46 short interfering RNA (siRNA) was transfected into ARPE-19 cells, and VEGF levels were determined by ELISA. Finally, in the same ECM conditions, ARPE-19 cells were challenged with normal human serum and VEGF levels determined by ELISA. RESULTS: CD46 is expressed on the basolateral surface of ARPE-19 cells on RPE-derived ECM. Nitrite modification of ECM reduced the expression of CD46 on ARPE-19 cells by 0.5-fold (P = 0.003) and increased VEGF release in ARPE-19 cells by 1.7-fold (P < 0.001). CD46 knockdown also increased release of VEGF on the apical and basal sides of ARPE-19 cells in culture by 1.3- (P = 0.012) and 1.2-fold (P = 0.017), respectively. CONCLUSIONS: Nitrite modification of the ECM decreased CD46 expression and increased the release of VEGF from ARPE-19 cells. Changes in CD46 expression may lead to changes in VEGF and play a pathologic role in the development of age-related macular degeneration.

The relationship between image degradation and myopia in the mammalian eye.

BACKGROUND: In all species studied, myopia develops if the eye is deprived of detailed vision during development (form deprivation myopia). However, different degrees of spatial image deprivation produce different effects and have not been described in the mammalian eye. Therefore, the effect of image degradation on guinea pig emmetropisation was investigated. METHODS: Eighty-one guinea pigs wore a treatment on one eye from 6 to 13 days of age. There were four treatments: a translucent diffuser (no lines or edges were visible through the diffuser); one of five Bangerter foils (BF: 0.8, 0.6, 0.4, 0.2, light perception only), which differed in their cut-off spatial frequencies; a 'ring mount' control with no filter; or one of two neutral density filters that reduced luminance only (ND, optical density grades 0.1 and 0.6). Refractive error and ocular elongation were measured after seven days of treatment. RESULTS: The extent of induced myopia and ocular growth were related to the amount of image degradation (mean difference between the treated and untreated eyes changed in a graded manner -7.0 D to -0.2 D and from 85 µm to seven µm respectively, for spatial frequency cut-offs between zero and 24 cycles per degree). Corresponding reductions in luminance from ND filters did not increase eye growth and caused significantly less myopia than the BFs that caused a similar luminance decrement. The greatest myopia occurred when no or limited spatial information was available to the eye, but moderate myopia still occurred with spatial frequency cut-offs of six and 12 cycles per degree, well beyond the visual acuity range of guinea pigs. CONCLUSION: Excessive ocular growth and myopia are most robust when induced by spatial frequency reductions within the visual acuity range but can also be induced beyond this. Either the mechanism of ocular growth can detect supra-threshold spatial frequencies, possibly due to aliasing, or it is sensitive to small amounts of contrast degradation.

Integration of defocus by dual power Fresnel lenses inhibits myopia in the mammalian eye.

PURPOSE: Eye growth compensates in opposite directions to single vision (SV) negative and positive lenses. We evaluated the response of the guinea pig eye to Fresnel-type lenses incorporating two different powers. METHODS: A total of 114 guinea pigs (10 groups with 9-14 in each) wore a lens over one eye and interocular differences in refractive error and ocular dimensions were measured in each of three experiments. First, the effects of three Fresnel designs with various diopter (D) combinations (-5D/0D; +5D/0D or -5D/+5D dual power) were compared to three SV lenses (-5D, +5D, or 0D). Second, the ratio of -5D and +5D power in a Fresnel lens was varied (50:50 compared with 60:40). Third, myopia was induced by 4 days of exposure to a SV -5D lens, which was then exchanged for a Fresnel lens (-5D/+5D) or one of two SV lenses (+5D or -5D) and ocular parameters tracked for a further 3 weeks. RESULTS: Dual power lenses induced an intermediate response between that to the two constituent powers (lenses +5D, +5D/0D, 0D, -5D/+5D, -5D/0D and -5D induced +2.1 D, +0.7 D, +0.1 D, -0.3 D, -1.6 D and -5.1 D in mean intraocular differences in refractive error, respectively), and changing the ratio of powers induced responses equal to their weighted average. In already myopic animals, continued treatment with SV negative lenses increased their myopia (from -3.3 D to -4.2 D), while switching to SV positive lenses or -5D/+5D Fresnel lenses reduced their myopia (by 2.9 D and 2.3 D, respectively). CONCLUSIONS: The mammalian eye integrates competing defocus to guide its refractive development and eye growth. Fresnel lenses, incorporating positive or plano power with negative power, can slow ocular growth, suggesting that such designs may control myopia progression in humans.

Temporal properties of the myopic response to defocus in the guinea pig.

PURPOSE: Hyperopic defocus induces myopia in all species tested and is believed to underlie the progression of human myopia. We determined the temporal properties of the effects of hyperopic defocus in a mammalian eye. METHODS: In Experiment 1, the rise and decay time of the responses elicited by hyperopic defocus were calculated in 111 guinea pigs by giving repeated episodes of monocular -4 D lens wear (from 5 to 6 days of age for 12 days) interspersed with various dark intervals. In Experiment 2, the decay time constant was calculated in 152 guinea pigs when repeated periods of monocular -5 D lens-wear (from 4 days of age for 7 days) were interrupted with free viewing periods of different lengths. At the end of the lens-wear period, ocular parameters were measured and time constants were calculated relative to the maximum response induced by continuous lens wear. RESULTS: When hyperopic defocus was experienced with dark intervals between episodes, the time required to induce 50% of the maximum achievable myopia and ocular elongation was at most 30 min. Saturated 1 h episodes took at least 22 h for refractive error and 31 h for ocular length, to decay to 50% of the maximum response. However, the decay was an order of magnitude faster when hyperopic defocus episodes were interrupted with a daily free viewing period, with only 36 min required to reduce relative myopia and ocular elongation by 50%. CONCLUSIONS: Hyperopic defocus causes myopia with brief exposures and is very long lasting in the absence of competing signals. However, this myopic response rapidly decays if interrupted by periods of 'normal viewing' at least 30 min in length, wherein ocular growth appears to be guided preferentially by the least amount of hyperopic defocus experienced.

The development of eye shape and the origin of lower field myopia in the guinea pig eye.

In a variety of species, the refractive state of the eye differs in different parts of the visual field (VF) with greater myopia in the region that views the ground ("lower field myopia"). We studied the refraction and eye shape of the normal guinea pig eye to determine what feature(s) underlie this visual adaptation. Guinea pigs (n=67) were either newborn or raised under incandescent light until 14, 37 or 45 days of age (20, 44, 20 and 11 eyes respectively). Refractive error was measured on-axis and 30° off-axis in the superior (SVF), inferior (IVF), temporal (TVF) and nasal (NVF) visual fields. Eye shape was analyzed from images of frozen hemisections in both the horizontal and vertical mid plane in 14 day animals, and in the vertical plane at 0, 14 and 45 days of age. Axial distances in vitro were correlated with in vivo high frequency ultrasound (r(2)=0.90). In the horizontal plane, asymmetry was caused by a ± 6° conical zone surrounding the optic nerve (12° off-axis in NVF), suggesting significant myopia in this zone. At 30°, there was no asymmetry in eye length, but the NVF was +1.7D more myopic due to asymmetry in corneal power. In the vertical plane at 30°, the IVF was more myopic than the SVF by -3.8D at 0 days, -5.9D at 14 days and -6.0D at 37 days. It resulted from vertical asymmetry in the distance of the retina from the lens center, which was longest in the mid IVF. This non-linear ramp retina was present at birth. In older animals, the peak of the ramp shifted more centrally, and the eye developed longer lengths in the extreme upper periphery (SVF) which may have been caused by the low position of the room ceiling. The vertical asymmetry in eye shape was mirrored by changes in choroid thickness, suggesting a mechanism by which eye shape was refined by vision during development. In early life, ocular growth in the vertical plane was 1.7 times higher in the center relative to the periphery, a pattern that reversed in the following month. Since emmetropization was achieved over this period, local visual cues related to clear vision may provide a switch to change ocular growth from a central to a peripheral emphasis.