Although visual circuit impairments in patients with AD patients has been reported at the macroscopic level , to the best of our knowledge, there is no study on the alteration of the visual circuit at the mesoscale level in the AD brain. Thus, our study aimed to provide direct anatomical evidence in a mouse model of AD by examining the topographical changes in various visual pathways of AD progression. To investigate the visual circuitry of the AD brain, we intravitreally injected a non-transsynaptic anterograde trace CTβ into the eyes of 4- and 12-month-old WT and 5XFAD mice. We demonstrated that the retinal projections to the SCN, dLGN, IGL, and vLGN reduced in the 4-month-old 5XFAD mice, which exhibit no cognitive decline, compared with the 4-month-old WT mice. In addition, we found decreased connections of retina-dLGN and retina-SC in the 12-month-old 5XFAD mice, representing severe stage of AD, compared to 12-month-old WT mice. Finally, we revealed that retinal connectivity decreases in several retinorecipient areas except for the OPN by normal aging. The present study demonstrated for the first time, the alteration of the visual pathways from the retina in the AD brain at the mesoscale level (Fig. 8).
It is well known that Aβ-induced neuronal dysfunctions induces neuronal circuit and network disturbances . A key component of these neuronal dysfunctions is axonal degeneration. Axonal degeneration by Aβ toxicity preceded neuronal cell death. In addition, inhibition of axonal degeneration through overexpression of nicotinamide mononucleotide adenylyltransferase 1 (Nmnat1) and Bcl-xl prevented neuronal cell death . This evidence suggests that Aβ targets the axon rather than the soma. Therefore, Aβ observed in the retina (Fig. 1f) can cause axonal degeneration of RGCs, resulting in disruption of the visual pathway. Damage to neuronal circuits leads to functional dysfunctions. Many visual-related dysfunctions have been reported in patients with AD and animal models [41, 42]. More than half of the patients with AD suffer from sleep disorders . Moreover, circadian rhythm disturbance was observed in 4-month-old 5XFAD mice . The accompanying circadian disruption in patients with AD and an animal model is closely related to the SCN, vLGN, and IGL [43, 44]. Surprisingly, the connections between the retina and the SCN, vLGN, and IGL were significantly impaired in the 4-month-old 5XFAD mice (Fig. 3c and 5d, e). These results suggest that the visual pathways in the SCN, vLGN, and IGL are severely impaired in the early-stage of AD, and the alteration of these circuitry results in abnormal circadian rhythms. In addition, it is well known that the dLGN is involved in the control of color vision and contrast sensitivity [45, 46]. Loss of contrast sensitivity and impairment of color vision is one of the early symptoms in patients with AD [47, 48]. Moreover, a study using an APPSWE/PS1∆E9 mouse model reported that contrast sensitivity and color vision were impaired in AD progression . Similarly, projection from the retina to the dLGN decreased in the 4- and 12-month-old 5XFAD mice (Fig. 5c, f). This finding indicated that the retina-dLGN pathway impaired AD progress, and the impairment of these circuitry resulted in the loss of contrast sensitivity and color vision. The SC is involved in visuomotor function, such as eye movement [23, 25]. Interestingly, visuomotor impairment was observed from 6 months of age in 5XFAD mice . Our results did not show the altered pathway from the retina to the SC in 4-month-old 5XFAD mice. In contrast, the retina-SC pathway was significantly reduced in 12-month-old 5XFAD mice (Fig. 6c, d). This finding demonstrates that retina-to-SC projections initiate impairment at the late stage of AD, and the disruption of retina-SC circuitry causes visuomotor dysfunction.
In the early-stage of Aβ-overexpressing mice, the circuits from the retina to SCN and LGN are impaired (Figs. 3 and 5), and the RGCs constituting these circuits are mainly ipRGCs . Specifically, M1 ipRGCs expressing the Brn3b transcription factor innervate into the dLGN and vLGN, and Brn3b-negative M1 ipRGCs project into the SCN and IGL . In addition, cadherin-6-expressing ipRGCs output to IGL and vLGN . Thus, these evidences suggest that Aβ-induced impairments of Brn3b-negative and positive M1 ipRGCs and their axons could be predominant in the early-stage of AD. In the late-stage of Aβ-overexpressing mice, pathways from the retina to the dLGN and SC are impaired (Fig. 5c and 6), these pathways consist of various RGCs axons . The dLGN receives inputs from a variety of RGCs, such as on, off, and on-off DSGCs and off alpha RGCs . Eighty to ninety percent of RGCs reach the SC . Based on several studies, most RGCs and their innervation could be impaired by various AD pathologies in the late-stage of AD.
Neurodegenerative biomarkers for detecting cognitive decline or MCI developing dementia is critical . One of the studies reports that ophthalmological examinations have revealed many visual alterations in individuals with neurodegenerative disease . Visual indications commonly precede brain symptoms in some disorders, suggesting that eye examinations could provide earlier detection of the underlying disease . Our results demonstrated that Aβ accumulation in the retina and impairment of visual pathways appear before the cognitive decline (Fig. 1f, 3d, and 5f). These results suggest that impaired retinas could be utilized to promptly diagnose AD in the early stage of the disease progression. Surprisingly, advances in ophthalmologic techniques for the study of the retina in vivo enable the diagnosis of AD through retinal degeneration. A non-invasive and inexpensive method of diagnosing AD through the retina can make a huge contribution to AD treatment. A recent review article presents several ocular biomarkers studied in patients with AD, such as decreased RNFL thickness, retinal vascular changes, and AD pathology detection in the retina . In addition to the diagnosis through retinal degeneration, various biomarkers exist that diagnose AD by measuring the visual dysfunction [55–58].
Our results suggested the retinal outputs were impaired in the 5XFAD mice. In particular, the most common neurotransmitter in the visual system, including the retina, is glutamate . Unfortunately, most conventional tracers are unable to reveal the connectivity at the level of the cell-type-specific pathway . Therefore, genetic tracing using cell-type-specific promoters should be used to investigate the disruption of glutamate-specific visual pathways. Nevertheless, the current research is the first to demonstrate the alteration in the visual pathways in an animal model of AD at the mesoscale level. Restoring damaged visual pathways with optogenetic stimulation, based on mesoscopic mapping of connectivity in the AD brain, could be a possible therapeutic approach for treating visual symptoms in patients with AD.
5XFAD mouse, an Aβ-overexpressing mouse model, has a high level of Aβ40 and Aβ42 peptide, which is the closest to humans in the retina and brain. Moreover, the 5XFAD mice showed stronger retinal and synaptic pathology than other AD mouse models . Significantly increased Aβ concentrations were observed in the eyes of young 5XFAD mice without cognitive impairment compared to WT mice of the same age [14, 62, 63]. In addition, visual behavior abnormalities and circadian rhythm disturbance were reported in 5XFAD mice [42, 50]. Unfortunately, 5XFAD mice do not reflect all of the pathologies of AD and may not reflect the same visual pathway impairment as patients with AD. Nevertheless, since non-clinical studies are needed for developing treatment and diagnostic methods for AD, among various AD models, we chose 5XFAD mice, which accumulate the Aβ in the retina at the early-stage and exhibit visual-related symptoms.