APOE4 expression confers a mild, persistent reduction in neurovascular function in the visual cortex and hippocampus of awake mice

Abstract

Vascular dysfunction is an early feature of late onset Alzheimer's disease (AD), preceding classic AD pathology such as beta amyloid accumulation and formation of hyperphosphorylated tau. Such vascular dysfunction may promote classic AD pathology by decreasing blood flow, impairing brain oxygenation and clearance of molecules such as beta amyloid. The main genetic risk factor for AD is the ϵ4 allele of APOE, which has been found to increase blood brain barrier permeability and decrease vascular density, as well as decrease blood flow and functional hyperaemia in anaesthetised mice undergoing acute surgery. These results suggest that APOE4 may confer AD risk via its effects on the vasculature. However, the responses of neurons and individual vessels have not been studied, so neurovascular relationships are unknown, and no previous studies have looked at awake mice. We therefore measured neurovascular responses at rest and in response to visual stimulation using 2 photon imaging of awake APOE3 and APOE4 targeted-replacement (APOE TR) mice that expressed the calcium indicator GCaMP6f in excitatory neurons, while labelling the vascular lumen with Texas Red dextran. In parallel, we measured cerebral blood flow, blood oxygenation and cerebral blood volume using combined laser Doppler flowmetry and haemoglobin spectrometry. Measurements were performed in mice aged between 3-4 months to 12-13 months. We found a milder vascular deficit in awake mice than previous studies that used an acute surgical preparation: capillary responses to visual stimulation were the same in APOE3 and APOE4 TR mice, leading to unimpaired functional hyperaemia. However, neuronal calcium signals during visual stimulation were significantly enhanced in APOE4 mice, while there was a marked decrease in pial arteriole responsiveness and vasomotion. This pattern of results was unaffected by age, suggesting that APOE4 expression creates a stable, but mildly altered neurovascular state that does not itself cause degeneration. However, these changes likely make the system more sensitive to subsequent insults; for example, weaker vasomotion could impair clearance of beta amyloid as it starts to accumulate, and therefore may help explain how APOE4 expression increases risk of developing AD.

Gradual Not Sudden Change: Multiple Sites of Functional Transition Across the Microvascular Bed

“…These recordings could be during rest, visual stimulus and/or locomotion, and we included the entire continuous trace in the power analysis as (a) the stimulus presentation occurred for 5 s every 30 s, meaning the power spectrum peak corresponding to stimulus presentation would be at 0.03 Hz (outside our 0.05-0.15 Hz range of interest); and (b) there was no significant difference in the amount of time spent in locomotion between the different vessel categories (mean time spent in locomotion: 13.07%, standard deviation: 6.39%, p = 0.45). All continuous data was detrended by subtracting the baseline (the 8th percentile calculated over a 15 s time window; see Bonnar et al, 2021 ). The MATLAB function “pwelch” was used to conduct discrete Fourier transforms across 60 s time windows.…”

“…Similar effects may be seen after stroke, with capillaries constricting in brain slices during ischemia ( Hall et al, 2014 ) and showing stalled flow and constrictions after reperfusion in vivo ( Yemisci et al, 2009 ), though the largest constrictions may be seen in αSMA-expressing vessels rather than mid-capillary regions ( Hill et al, 2015 ). Conversely, in mice carrying the main genetic risk factor for Alzheimer’s disease, APOE4, capillary function was unaffected, but the large pial vessels were dysfunctional, with vasomotion and dilation frequencies reduced compared to APOE3-expressing controls ( Bonnar et al, 2021 ). However, none of these studies define the precise location of these transitions in disease susceptibility.…”

“…In a mouse model of Alzheimer’s disease, arterioles surrounded with Aβ showed impaired vasomotion when driven by neuronal activity, and showed reduced dextran clearance from the parenchyma ( van Veluw et al, 2020 ). Mice carrying the main genetic risk factor for Alzheimer’s disease, APOE4, also showed a reduction in pial arteriole vasomotion compared to APOE3 controls ( Bonnar et al, 2021 ). Thus, vasomotion is important for regulating supply and clearance of substances to the brain, and is affected by disease.…”