Introduction:
Cerebral amyloid angiopathy (CAA), the accumulation of amyloid-
β
(1-40)
(A
β
) around cerebral arteries, impairs endothelial function. Endothelium-dependent dilation is a consequence of transient increases in intracellular [Ca
2+
] in endothelial cells (EC). The Ca
2+
permeable N-methyl-D-aspartate receptor (NMDAR) mediates endothelium-dependent dilation, although if these effects are dependent on Ca
2+
influx and transients, or if they are impaired by A
β
, remains undetermined.
Hypothesis:
A
β
inhibits endothelial NMDAR-mediated Ca
2+
influx and transients in murine pial arteries.
Methods:
We performed Ca
2+
time-lapse imaging of
en face
pial arteries from
cdh5-GCaMP8
mice to quantify EC Ca
2+
events induced by NMDAR activation. Data are means ± SEM.
Results:
Elemental Ca
2+
entry through NMDAR, hereon called
NMDAR sparklets
, was assessed in arteries incubated with EGTA-AM and cyclopiazonic acid (CPA) to inhibit intracellular Ca
2+
transients. NMDA (10 μM) induced an increase in
NMDAR sparklets
frequency when compared to vehicle, an effect inhibited by the NMDAR antagonist D-APV (in Hz: 0.12±0.01 vs 0.44±0.05 vs 0.21±0.02, vehicle vs NMDA vs NMDA+D-APV, p<0.05). Further, pial arteries exposed to NMDA without EGTA-AM and CPA showed an increase in the frequency of intracellular Ca
2+
transients, also blocked by D-APV (in Hz: 0.24±0.05 vs 0.53±0.10 vs 0.28±0.05, vehicle vs NMDA vs NMDA+D-APV, p<0.05). We then tested the effects of A
β
on Ca
2+
events in pial artery EC. We observed that 30 minutes exposure to A
β
(5 μM) caused a significant reduction in
NMDAR sparklets
(in Hz: 0.62±0.07 vs 0.22±0.03, NMDA vs NMDA + A
β
, p<0.05) but did not significantly alter intracellular Ca
2+
transients (in Hz: 0.62±0.37 vs 0.27±0.07, NMDA vs NMDA + A
β
). Lastly, we performed pressure myography on pial arteries of wild-type and
5x-FAD
mice, a model of familial Alzheimer’s disease with rapid amyloid accumulation.
5x-FAD
mice displayed impaired vasodilation to NMDA (vasodilation (%): 9.86±0.64 vs 4.22±2.76, wild-type vs
5x-FAD
, p<0.05).
Conclusion:
These preliminary data suggest that A
β
impairs endothelial NMDAR-associated Ca
2+
influx events in cerebral arteries, which can impair blood flow in CAA patients, thus contributing to cognitive impairment.