Amyloid β alters vascular Ca_V1.2 channel spatiotemporal properties

A hallmark of Alzheimer's disease is reduced cerebral blood flow, which appears to occur early in disease progression and has been associated with amyloid β (Aβ) accumulation. Cerebral artery diameter is regulated by calcium influx via voltage-gated L-type Ca_V1.2 channels, which contribute to arterial myocyte contractility. The potential effects of Aβ on vascular Ca_V1.2 channels remain unclear. To address this knowledge gap, we test the hypothesis that Aβ alters vascular Ca_V1.2 channel spatiotemporal properties. Using freshly dissected cerebral arteries and arterial myocytes from wild-type (WT) mice, super-resolution imaging revealed that Aβ_1-42, but not Aβ_1-40, increased the clustering of Ca_V1.2 channels in male but not female or ovariectomized female arterial myocytes. This Aβ_1-42-associated Ca_V1.2 clustering appears to depend on NADPH oxidases, protein kinase C and protein kinase A signalling. Studies using male arterial myocytes from S1928A knockin mice suggested that Aβ_1-42 effects on Ca_V1.2 clustering require phosphorylation of the α1_C/Ca_V1.2 pore-forming subunit at serine-1928. Single-channel electrophysiology showed increased Ca_V1.2 channel activity and cooperative gating in WT male arterial myocytes exposed to Aβ_1-42, but not in female WT or male S1928A myocytes. In functional studies, WT, but not S1928A, cerebral arteries acutely incubated with Aβ_1-42 demonstrated enhanced constriction in response to the Ca_V1.2 channel agonist BayK8644. These findings provide insight into mechanisms by which Aβ may influence Ca_V1.2 properties and vascular contractility, potentially contributing to vascular changes in Alzheimer's disease. KEY POINTS: Specific amyloid-β peptide (Aβ_1-42) induces α1_C/Ca_V1.2 super-clustering in cerebral vascular smooth muscle, enhancing Ca_V1.2 activity. Aβ_1-42 engages distinct pathways involving NOX-derived ROS-activated PKC and PKA (through unknown mechanisms) to drive α1_C/Ca_V1.2 spatiotemporal reorganization leading to enhance cerebral artery contractile responses to Ca_V1.2 agonists. The Aβ_1-42-mediated PKC- and PKA-dependent effects on α1_C/Ca_V1.2 spatiotemporal remodelling and cerebral artery contractility require phosphorylation of α1_C at S1928, as the effects are lost in S1928A tissue/cells. The molecular and functional changes occur exclusively in male vascular smooth muscle cells, revealing sex-specific vulnerabilities in amyloid β-related cerebrovascular pathology that are independent of sex hormones and establishing a potential mechanism for vascular dysfunction in conditions with elevated Aβ_1-42.