Endothelial Arf6 sustains electrical signaling and cerebral blood flow in mice through PIP2-dependent activation of Kir2.1 channels.

Brain capillaries sense neural activity and direct blood flow to active regions-a process termed neurovascular coupling that underlies activity-dependent increases in local perfusion (functional hyperemia). A key contributor to functional hyperemic responses is the capillary endothelial cell (cEC) inward rectifier K+ (Kir2.1) channel, which, when activated by neuronal activity-derived extracellular K+, initiates vasodilatory electrical signals that propagate through the vascular network. Kir2.1 channel function requires continual production of its lipid cofactor, phosphatidylinositol-4,5-bisphosphate (PIP2), and is compromised in mouse models of cerebral small vessel (cSVD). Although decreased PIP2 availability is a common feature of cSVDs, mechanisms underlying PIP2 synthesis remain poorly understood. We hypothesized that Arf6, a small GTPase expressed in cECs that stimulates PIP2 production, is critical for this process. Using patch-clamp electrophysiology, we demonstrate that inhibiting Arf6 activity progressively decreased cEC Kir2.1 channel activity. This deficit manifested as loss of capillary-to-arteriole electrical signaling in isolated vessels and diminished functional hyperemia in vivo. Exogenously provided PIP2 restored Kir2.1 currents and functional hyperemia after Arf6 inhibition or genetic knockdown. Collectively, our data suggest that cEC Arf6 sustains Kir2.1 activity by maintaining PIP2 levels and demonstrate that diminished PIP2 synthesis is sufficient to impair functional hyperemia. Furthermore, we identify Arf6 as a mechanistic link between PIP2 production and endothelial electrical signaling, highlighting Arf6 as a potential therapeutic target for restoring functional hyperemia.