Cognitive decline is linked to decreased cerebral blood flow, particularly in women after menopause. Impaired cerebrovascular function precedes the onset of dementia, possibly due to reduced functional dilation in parenchymal arterioles. These vessels are bottlenecks of the cerebral microcirculation, and dysfunction can limit functional hyperemia in the brain. Large conductance Ca2+-activated K+ channels (BKCa) are the final effectors of several pathways responsible for functional hyperemia, and their expression is modulated by estrogen. However, it remains unknown if BKCa function is altered in cerebral parenchymal arterioles after menopause. Using a chemically-induced model of menopause, the 4-vinylcyclohexene diepoxide (VCD) model, which depletes follicles while maintaining intact ovaries, we hypothesized that menopause would be associated with reduced functional vasodilatory responses in cerebral parenchymal arterioles of wild-type mice via reduced BKCa function. Using pressure myography of isolated parenchymal arterioles, we observed that menopause (Meno) induced a significant increase in spontaneous myogenic tone. Endothelial function, assessed as nitric oxide production and dilation after cholinergic stimulation or endothelium-dependent hyperpolarization pathways, was unaffected by Meno. BKCa function was significantly impaired in Meno compared to Control, without changes in voltage-gated K+ channel activity. Cerebral functional hyperemia, measured by laser speckle contrast imaging during whiskers stimulation, was significantly blunted in Meno mice, without detectable changes in basal perfusion. However, behavioral testing identified no change in cognition. These findings suggest that menopause induces cerebral microvascular and neurovascular deficits.

Vasculopathy and nitro-oxidative stress are present in patients with Alzheimer’s disease (AD) and may contribute to disease progression and severity. Large conductance calcium activated K+ channels (BKCa) plays an important role in vasodilatory responses and maintenance of myogenic tone in resistance arteries. Opening of BKCa channels occurs upstream from localized intracellular Ca2+ release events (Ca2+ sparks), and results in K+ efflux, vascular smooth muscle cell hyperpolarization and vasorelaxation. In a pro-nitro-oxidative scenario, BKCa can be modified, resulting in decreased activity and hypercontractility, which can compromise cerebral blood flow regulation, generating an environment that may accelerate neurodegeneration. We hypothesized that reductions in BKCa-dependent vasodilation in cerebral arteries, as consequence of nitro-oxidative stress, results in neurovascular dysfunction in the 5x-FAD model of AD. Methods: Posterior communicating arteries (PComA) from 5 months-old male and female 5x-FAD and wild-type (WT) littermates were isolated and studied ex vivo using pressure myography. Smooth muscle Ca2+ transients were evaluated by spinning-disk confocal microscopy. Oxidative stress was assessed by oxidized glutathione levels in the brain using a colorimetric enzymatic assay. BKCa expression was assessed by qPCR. Nitrosylated BKCa was evaluated using Western blot. Functional hyperemia were evaluated by laser speckle contrast imaging. Data are means±SEM, 5x-FAD vs WT, analyzed by two-tailed Student’s t-test or Mann-Whitney test. Results: In females, PComA from 5x-FAD showed higher spontaneous myogenic tone than WT (Myogenic tone: 24.48 ± 3.20 vs 16.09 ± 0.93%, p<0.05, N=7). Constriction to the BKCa blocker iberiotoxin (30 nM) was smaller in 5x-FAD than WT, suggesting lower basal BKCa activity (Vasoconstriction: -4.25 ± 0.43 vs -9.22 ± 2.56%, p<0.05; N=5), which was independent of alterations in intracellular Ca2+ transients or BKCa mRNA expression. These vascular changes were associated with higher levels of oxidized glutathione in female 5x-FAD (7.83 ± 0.62 vs 5.27 ± 0.74 μM, p<0.05, N=8) and of S-nitrosylation in the BKCa α-subunit (0.68 ± 0.04 vs 0.41 ± 0.03, p<0.05, N=5). Female 5x-FAD mice showed increased expression of iNOS mRNA ([2-ΔΔCT]: 10.64 ± 5.40 vs 0.74 ± 0.19, p<0.05, N=6) and impaired functional hyperemia responses after whisker stimulation (%increase: 3.82 ± 0.64 vs. 9.91 ± 1.41%, p<0.05, N=6). No significant differences were observed between male 5x-FAD and WT for all parameters above. Conclusion: Cerebrovascular impairments were more pronounced in female 5x-FAD mice, observed as an increase in spontaneous myogenic tone, likely due to reduction in smooth muscle cell BKCa activity associated to an increase in brain nitro-oxidative stress and a blunted neurovascular coupling response. Together, they identify post-translational modifications of BKCa as a putative target to improve cerebral microvascular function in AD. National Institutes of Health (R00 HL140106, R01 AG073230) and the Alzheimer's Association (AARGD-21-850835). This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Cerebral microvascular dysfunction and nitro-oxidative stress are present in patients with Alzheimer's disease (AD) and may contribute to disease progression and severity. Large conductance Ca2+-activated K+channels (BKCa) play an essential role in vasodilatory responses and maintenance of myogenic tone in resistance arteries. BKCacan be modified in a pro-nitro-oxidative environment, resulting in decreased activity and vascular hyper-contractility, which can compromise cerebral blood flow regulation. We hypothesized that reductions in BKCafunction in cerebral arteries, as a consequence of nitro-oxidative stress, are associated with blunted neurovascular responses in the5x-FADmodel of AD. Using pressure myography, we observed that posterior communicating arteries (PComA) from 5 months-old female5x-FADmice showed higher spontaneous myogenic tone than wild-type (WT) littermates. Constriction to the BKCablocker iberiotoxin (30 ηM) was smaller in5x-FADthan WT, suggesting lower basal BKCaactivity, which was independent of alterations in intracellular Ca2+transients or BKCamRNA expression. These vascular changes were associated with higher levels of oxidative stress in female5x-FADand a higher level of S-nitrosylation in the BKCaα-subunit. In females, pre-incubation of PComA from5x-FADwith the reducing agent DTT (10 μM) rescued iberiotoxin-induced contraction. Female5x-FADmice showed increased expression of iNOS mRNA, lower resting cortical perfusion atop the frontal cortex, and impaired neurovascular coupling responses. No significant differences between male5x-FADand WT were observed for all parameters above. These data suggest that the exacerbation in BKCaS-nitrosylation contributes to cerebrovascular and neurovascular impairments in female5x-FADmice.