Using chromatic pupilloperimetry testing in conjunction with machine learning algorithms we demonstrated that multi-parametric analysis of PLR kinetics for focal red and blue light stimuli can differentiate with high discriminative power cognitively asymptomatic individuals at high risk for AD due to a parental family history from individuals with no family history of AD.
The main contributions of this study are two; First, our prediction model could classify normally cognitively asymptomatic individuals according to the existence of AD family history, which is a major risk factor for developing the disease. Second, the prediction model achieving highest AUC-ROC was based on parameters pertaining to the latency of the transient PLR contraction phase. Our findings are supported by studies demonstrating changes in AD patients compared to healthy controls in PLR parameters related to the afferent arm of the reflex. Among them, some studies reported changes in latency-related PLR parameters 21,22, while others found changes in PLR parameters pertaining to the contraction phase, but not to the latency of the response 16,17,23. To the best of our knowledge this is the first study that shows an association of attenuation of the afferent PLR arm with AD risk due to a parental family history in asymptomatic middle-aged adults. Our machine learning algorithm may detect changes in PLR in individuals at high risk for AD, years before the onset of cognitive decline.
The PLR parameter that achieved the highest performance rates by our learning algorithm for both eyes was the latency of the transient PLR, suggesting an effect on the contraction arm of the PLR, which is attributed to parasympathetic system. This finding is supported by studies demonstrating that AD Patients have reduced synthesis of Acetylcholine 39, the key neurotransmitter in the para-sympathetic nervous system 40. AD affects the cholinergic system 41, and specifically the cholinergic Edinger Westphal Nucleus (EWN), 42,43 which is involved in the control of the pupil contraction 44. Our findings are in accordance with published studies with patients with probable AD reporting attenuated pupil contraction acceleration 22,45,46, shorter PLR latency and lower amplitude of pupil contraction 45.
Using the pupil response latency (PRL) parameter, the highest performance values were obtained for both eyes were in response to dim blue light with lower, yet still very high performance for red light. These results demonstrate a slightly higher performance for the blue light illumination indicating that attenuation of the rod PLR pathway may be a very early event in the pre-AD stage. Interestingly, the most discriminative test targets of a positive AD family history in response to dim blue light were located in periphery of the visual field, in the temporal area, whereas in response to dim red light stimuli, discriminative test targets were detected across the visual field with no specific pattern. These finding may suggest attenuated function of the neuronal circuits downstream of rods in the nasal retina in early stages of AD pathogenesis. Furthermore, these findings indicate the possibility of running a shorter chromatic pupilloperimetry test, using a smaller number of test targets spots in the temporal visual field, that may suffice for screening using such classification algorithm. This should be validated in further studies. Differences in rod pathway function were also observed in full-field electrophysiology studies in mouse models of AD, where abnormal dark adapted retinal function was recorded, whereas cone photoreceptor function and signal transmission to second order neurons was maintained 47,48.
Since the PLR for bright red and blue light had low discrimination between groups, our data suggest that at the early pre-AD stage, the extrinsic ipRGC activation is affected whereas the melanopsin-mediated intrinsic activation of ipRGCs remains intact. Our findings are in accordance with the finding of Romagnoli et al who reported similar melanopsin-mediated but aberrant rod-mediated PLR in mild-moderate AD patients using full-field chromatic pupillometry 33. Moreover, a recent pathology postmortem study indicated significant thinning of ipRGC dendrites in retinas of AD patients13, providing a structural support for our functional findings.
All parameters with AUC-ROC > 0.7 were related to PLR latency suggesting a possible attenuation of the speed of neuronal signaling associated with high risk of AD disease. The only latency-related parameter that had AUC-ROC <0.7 for both eyes was the latency of maximal velocity (LMCV). Several studies indicated that the LMCV parameter differentiates with high specificity and sensitivity between healthy controls and patients with photoreceptor degeneration35,37. SD-OCT imaging for all participants included in the current study did not indicate any sign of photoreceptor degeneration. Hence, our findings that the LMCV did not change in response to either red or blue light, while other latency parameters differentiated between the study groups, suggest that the PLR pathology at the pre-clinical stage of AD does not occur at the photoreceptor cell level but rather downstream at the bipolar/amacrine or ipRGC dendrites. These findings are in accordance with the findings of McAnany et al., who have most recently reported post-receptor abnormalities in retinal function in a mouse model of AD under dark adaptation using fill field ERG 47.
Our study has several limitations. First, our cohort included 186 participants. Since each participant had PLR examinations done on both eyes, we ran our experiments on two datasets, a dataset for the left eye and for the right eye, with each dataset consisting of data extracted from 372 eyes. Although our data set was rather large, verification of the results in larger cohorts may assist in strengthening our conclusions. In addition, our study was cross sectional. Hence, we can only speculate that pupil function alterations predict the development of AD, but this must be tested in a longitudinal design, which is ongoing in the IRAP study 49. Another limitation of our study is that it included only Caucasian subjects, which are the majority in the Israeli population. Hence, future studies including non-Caucasian individuals are required to expand the generalizability of our findings.
A third limitation concerns the ratio between subjects with and without AD family history. Most participants in our study had a confirmed AD family history, and they accounted for 67.2% of the study's participants. This number does not reflect the normal proportion of this condition in the population. This is attributed to the fact that our study participants are part of the IRAP cohort, which was formed as part of a longitudinal study 49. Participants are required to undergo a battery of clinical testing including cognitive assessment, MRI and blood tests every several years. In this type of study design recruiting healthy subjects in more challenging.
To the best of our knowledge, this is the first attempt to identify focal changes in the rod- cone and melanopsin-mediated PLR in cognitively normal, asymptomatic participants at high risk for AD. Using chromatic pupilloperimetry testing in a relatively large cohort and artificial intelligence tools, our study suggests that individuals at high risk to develop AD have specific changes, already in midlife, in the latency of PLR for focal dim red and blue light stimuli. Rod-circuits are specifically affected at the nasal retina whereas the cone-circuits are affected across the central retina, and melanopsin function remains unaffected at this early stage.