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Chronic inflammation is a constitutive component of many age-related diseases, including age-related macular degeneration (AMD). Here, we identified interleukin-1 receptor–associated kinase M (IRAK-M) as a key immunoregulator in retinal pigment epithelium (RPE) that declines during the aging process. Rare genetic variants of IRAK3, which encodes IRAK-M, were associated with an increased likelihood of developing AMD. In human samples and mouse models, IRAK-M abundance in the RPE declined with advancing age or exposure to oxidative stress and was further reduced in AMD. Irak3-knockout mice exhibited an increased incidence of outer retinal degeneration at earlier ages, which was further exacerbated by oxidative stressors. The absence of IRAK-M led to a disruption in RPE cell homeostasis, characterized by compromised mitochondrial function, cellular senescence, and aberrant cytokine production. IRAK-M overexpression protected RPE cells against oxidative or immune stressors. Subretinal delivery of adeno-associated virus (AAV)–expressing human IRAK3rescued light-induced outer retinal degeneration in wild-type mice and attenuated age-related spontaneous retinal degeneration in Irak3-knockout mice. Our data show that replenishment of IRAK-M in the RPE may redress dysregulated pro-inflammatory processes in AMD, suggesting a potential treatment for retinal degeneration.

Diabetic retinopathy is the most common microvascular complication of diabetes and is a major cause of blindness, but an understanding of the pathogenesis of the disease has been hampered by a lack of accurate animal models. Here, we explore the dynamics of retinal cellular changes in the Nile rat (Arvicanthis niloticus), a carbohydrate-sensitive model for type 2 diabetes. The early retinal changes in diabetic Nile rats included increased acellular capillaries and loss of pericytes that correlated linearly with the duration of diabetes. These vascular changes occurred in the presence of microglial infiltration but in the absence of retinal ganglion cell loss. After a prolonged duration of diabetes, the Nile rat also exhibits a spectrum of retinal lesions commonly seen in the human condition including vascular leakage, capillary non-perfusion, and neovascularization. Our longitudinal study documents a range and progression of retinal lesions in the diabetic Nile rat remarkably similar to those observed in human diabetic retinopathy, and suggests that this model will be valuable in identifying new therapeutic strategies.

Diabetic retinopathy is the most common microvascular complication of diabetes and is a major cause of blindness, but an understanding of the pathogenesis of the disease has been hampered by a lack of accurate animal models. Here, we explore the dynamics of retinal cellular changes in the Nile rat (Arvicanthis niloticus), a carbohydrate-sensitive model for type 2 diabetes. The early retinal changes in diabetic Nile rats included increased acellular capillaries and loss of pericytes that correlated linearly with the duration of diabetes. These vascular changes occurred in the presence of microglial infiltration but in the absence of retinal ganglion cell loss. After a prolonged duration of diabetes, the Nile rat also exhibits a spectrum of retinal lesions commonly seen in the human condition including vascular leakage, capillary non-perfusion, and neovascularization. Our longitudinal study documents a range and progression of retinal lesions in the diabetic Nile rat remarkably similar to those observed in human diabetic retinopathy, and suggests that this model will be valuable in identifying new therapeutic strategies.

Leber congenital amaurosis (LCA) is an inherited retinal dystrophy that causes childhood blindness. Photoreceptors are especially sensitive to an intronic mutation in the cilia-related gene CEP290, which causes missplicing and premature termination, but the basis of this sensitivity is unclear. Here, we generated differentiated photoreceptors in three-dimensional optic cups and retinal pigment epithelium (RPE) from iPSCs with this common CEP290 mutation to investigate disease mechanisms and evaluate candidate therapies. iPSCs differentiated normally into RPE and optic cups, despite abnormal CEP290 splicing and cilia defects. The highest levels of aberrant splicing and cilia defects were observed in optic cups, explaining the retinal-specific manifestation of this CEP290 mutation. Treating optic cups with an antisense morpholino effectively blocked aberrant splicing and restored expression of full-length CEP290, restoring normal cilia-based protein trafficking. These results provide a mechanistic understanding of the retina-specific phenotypes in CEP290 LCA patients and potential strategies for therapeutic intervention.

This paper documents a research project investigating the effects of various driving-while-intoxicated (DWI) checkpoint setup scenarios on overall traffic operations. According to NHTSA, 13,470 people were killed in alcohol-related crashes in 2006. Of the 13,470 fatalities, 64% involved drivers with a blood alcohol content of 0.08 or higher. For this reason, state and local police departments tend to conduct DWI checkpoints during the night, when police officers detain vehicles traveling on a roadway and interview drivers to determine suspicion of alcohol use. For planning and budgeting purposes, police departments need to be able to effectively plan a setup for DWI checkpoints. The goal of the project is to provide police departments with data and methodologies to analyze the impact of their checkpoints on traffic operations. Seven scenarios were investigated on the basis of interviews with several police departments: one officer stopping every vehicle, one officer stopping every second vehicle, one officer stopping every fifth vehicle, one officer stopping every tenth vehicle, two officers stopping every vehicle, four officers stopping every vehicle, and four officers stopping every second vehicle. Each scenario was varied to include 50, 100, 150, 200, 300, 400, 500, 600, 700, and 800 vehicles per hour (or vehicles per hour per lane) to obtain a wide range of limiting values after which DWI checkpoints would cause extraordinary amounts of delay. The paper includes the results for each scenario that can be used by police departments to recognize the potential impacts on vehicle delay.

Vascular pathologies are known to be associated with age-related macular degeneration. Recently, age-related macular degeneration was associated with a single-nucleotide substitution of the complement factor H (CFH) gene, part of the alternative pathway of the complement system, a critical element in the innate immune response. Such polymorphisms are found in more than 50% of cases of age-related macular degeneration. Here we show that the absence of CFH causes an autoimmune response that targets the vascular endothelium of both the inner and outer retinal vascular networks. In CFH-knockout (cfh−/−) mice, C3 and C3b, key components of the complement system, are progressively deposited on retinal vessels, which subsequently become restricted and wither, resulting in a reduction of retinal blood supply. This result leads to increased oxygen stress. While such effects are not systemic, these structural changes are mirrored in functional changes with a substantial decline in retinal blood flow dynamics. When the system is challenged functionally by laser-induced choroidal neovascularization, fluorescein leakage was significantly smaller in cfh−/−mice compared with controls, likely due to reduced retinal perfusion. These data reveal that in both the presence and absence of exogenous challenge to the innate immune system, CFH is required to maintain normal levels of retinal perfusion. It is likely that C3 and C3b accumulation in the aged CFH-deficient retina is associated with complement-mediated retinal endothelium destruction.

This paper documents a research project investigating the effects of various driving-while-intoxicated (DWI) checkpoint setup scenarios on overall traffic operations. According to NHTSA, 13,470 people were killed in alcohol-related crashes in 2006. Of the 13,470 fatalities, 64% involved drivers with a blood alcohol content of 0.08 or higher. For this reason, state and local police departments tend to conduct DWI checkpoints during the night, when police officers detain vehicles traveling on a roadway and interview drivers to determine suspicion of alcohol use. For planning and budgeting purposes, police departments need to be able to effectively plan a setup for DWI checkpoints. The goal of the project is to provide police departments with data and methodologies to analyze the impact of their checkpoints on traffic operations. Seven scenarios were investigated on the basis of interviews with several police departments: one officer stopping every vehicle, one officer stopping every second vehicle, one officer stopping every fifth vehicle, one officer stopping every tenth vehicle, two officers stopping every vehicle, four officers stopping every vehicle, and four officers stopping every second vehicle. Each scenario was varied to include 50, 100, 150, 200, 300, 400, 500, 600, 700, and 800 vehicles per hour (or vehicles per hour per lane) to obtain a wide range of limiting values after which DWI checkpoints would cause extraordinary amounts of delay. The paper includes the results for each scenario that can be used by police departments to recognize the potential impacts on vehicle delay.

Photoreceptor survival in the dystrophic rat was evaluated following administration of IL-1β at dosages much lower than those used previously for this purpose. Royal College of Surgeons rats (pink-eyed, pigmented, or non-dystrophic) received 1μl intravitreal injections of murine recombinant IL-1β (0.5, 2, or 5μgml−1; at 3 or 4 weeks of age). Eyes were harvested 4 weeks later and outer nuclear layer profiles counted. Additional animals received intravitreal basic fibroblast growth factor (1000μgml−1), or vehicle alone. Others were treated with IL-1β to evaluate the inflammatory response (CD45+ profiles) or visual function via opto-kinetic response. IL-1β was associated with photoreceptor rescue that was both dose-dependent and comparable to that seen following high-dose basic fibroblast growth factor. Significant anatomical rescue relative to controls was seen in both pink-eyed and pigmented strains, although the degree and distribution varied between strains. Functional rescue was confirmed by opto-kinetic response using the pigmented strain. At 5μgml−1, IL-1β resulted in numerous CD45+ profiles within the retina and vitreous. Infiltration peaked at 48hr and was minimal at 4 weeks, without dysplastic sequelae. IL-1β therefore induces visually significant photoreceptor rescue in a potent, dose-dependent manner that need not entail cytoarchitectural disruption. This is consistent with the known association between injury and rescue in the rat retina. Neuroprotection may be a general, if under-appreciated, consequence of inflammatory cascade activation.

This research investigates the hemodynamic response to stimulation of the barrel cortex in anaesthetized rats using optical imaging and spectroscopy (Bonhoeffer and Grinvald, 1996; Malonek and Grinvald, 1996; Mayhew et al.,1999). A slit spectrograph was used to collect spectral image data sequences. These were analyzed using an algorithm that corrects for the wavelength dependency in the optical path lengths produced by the light scattering properties of tissue. The analysis produced the changes in the oxy- and deoxygenation of hemoglobin following stimulation. Two methods of stimulation were used. One method mechanically vibrated a single whisker, the other electrically stimulated the whisker pad. The electrical stimulation intensity varied from 0.4 to 1.6 mA. The hemodynamic responses to stimulation increased as a function of intensity. At 0.4 mA they were commensurate with those from the mechanical stimulation; however, the responses at the higher levels were greater by a factor of ∼10. For both methods of data collection, the results of the spectroscopic analysis showed an early increase in deoxygenated hemoglobin (Hbr) with no evidence for a corresponding decrease in oxygenated hemoglobin (HbO2). Evidence for increased oxygen consumption (CMRO2) was obtained by converting the fractional changes in blood volume (Hbt) into estimates of changes in blood flow (Grubb et al.,1974) and using the resulting time course to scale the fractional changes in Hbr. The results show an early increase CMRO2peaking ∼2 s after stimulation onset. Using these methods, we find evidence for increased oxygen consumption following increased neural activity even at low levels of stimulation intensity.

Imaging of neural activation has been used to produce maps of functional architecture and metabolic activity. There is some uncertainty associated with the sources underlying the intrinsic signals. It has been reported that following increased neural activity there was little increased oxygen consumption (∼5%), although glucose consumption increased by ∼50%. The research we describe uses a modification of the Beer–Lambert Law called path-length scaling analysis (PLSA) to analyze the spectra of the hemodynamic and metabolic responses to vibrissal stimulation in rat somatosensory cortex. The results of the PLSA algorithm were compared with those obtained using a linear spectrographic analysis method (we refer to this as LMCA). There are differences in the results of the analysis depending on which of the two algorithms (PLSA or LMCA) is used. Using the LMCA algorithm, we obtain results showing an increase in the volume of Hbr at ∼2 s, following onset of stimulation but no complementary decrease in oxygenated haemoglobin (HbO2). These results are similar to a previous report. In contrast, after using the PLSA algorithm, the time series of the chromophore changes shows no evidence for an increase in the volume of deoxygenated haemoglobin (Hbr). However, after further analysis of the time series from the PLSA using general linear models (GLM) to remove contributions from low frequency baseline oscillations, both the HbO2and Hbr times series of the response to stimulation were found to be biphasic with an early decrease in saturation peaking ∼1 s after onset of stimulation followed by a larger increase in saturation peaking at ∼3 s. Finally, following the PLSA-then-GLM analysis procedure, we do not find convincing evidence for an increase in cytochrome oxidation following stimulation, though we demonstrate the PLSA algorithm to be capable of disassociating changes in cytochrome oxidation state from changes in hemoglobin oxygenation.

Ambient sulfates, total suspended particulates (TSP), and ozone data were collected in rural areas of New York and New Jersey as part of a study conducted jointly by the New York State Department of Environmental Conservation and the Interstate Sanitation Commission during July and August 1975. Relationships between these contaminants and various meteorological systems were examined. The sulfate and TSP data were obtained from 24-hour integrated high volume samples obtained on a daily basis at seven rural sites with the emphasis in this paper being on two sites: Whiteface Mountain, N.Y., and High Point, N.J. The sulfate concentrations were found to be highest during four distinct periods which were generally defined by anticyclonic stagnation and/or return flow of maritime tropical air. The highest values recorded at Whiteface Mountain and High Point were 24 μg/m3 and 20 μg/m3, respectively, in the period from July 21 through 25. During the other three periods, a maximum of at least 15 μg/m3 was observed at one or both sites. Coincidence of the high level periods at the two sites separated by approximately 200 miles and at five other rural New York State sites during most of the study is a strong indication of long-range transport. -The maximum sulfate concentration recorded during the study was 38 μg/m3 at Schoharie, N.Y.The SO4/TSP ratio (maximum 0.53) at High Point and Whiteface Mountain was increased, though nonuniformily, with higher TSP during the episode periods. Ozone concentrations at High Point were increased simultaneously with sulfates, as in previous observations at Whiteface Mountain1 which suggests it is either an important reactant in the formation of sulfates or these contaminants are concommitant in photochemical processes and/or summertime anticyclonic systems.

Sequences of images of the cortical surface can be processed to reveal information about the cortical microcirculation, regional cerebral blood flow (rCBF), and changes induced by neuronal activity. This study examined the use of different analysis methodologies on intrinsic optical images taken from rat sensory motor cortex and testes. Generalized linear model (GLM) analysis was used and compared with standard signal processing methods including principal component analysis. The GLM method has been used by Fristonet al.(1994,Hum. Brain Map.,1: 214–220) in the analysis of functional magnetic resonance imagery to identify regions of focal activity. We investigated the use of this method to analyze video image data of the modulation of rCBF from rat cortex. The results revealed spatiotemporal variations in rCBF in response to stimulation within local regions of cortex. The advantage of the GLM method is that it augments ordinary signal processing methods with an estimate of statistical reliability. The use of different wavelengths of illumination reveals spatial structures with different temporal relationships. In image time series data collected under green and red illumination a phase difference was found in the low frequency ≈0.1 Hz vasomotion oscillation. This phase difference occurred in data from both cortex and testes. A possible explanation of these differences is that the spectral absorption characteristics of the tissue reflect changes in the volume proportions of the different hemoglobin derivatives in interacting with the modulation of the volume of blood. It is suggested that the combination of these effects produces the phase differences we detect.

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