Shaun Sandow

Coupling red blood cell (RBC) supply to O 2 demand is an intricate process requiring O 2 sensing, generation of a stimulus, and signal transduction that alters upstream arteriolar tone. Although actively debated, this process has been theorized to be induced by hypoxia and to involve activation of endothelial inwardly rectifying K + channels (K IR ) 2.1 by elevated extracellular K + to trigger conducted hyperpolarization via connexin40 (Cx40) gap junctions to upstream resistors. This concept was tested in resting healthy skeletal muscle of Cx40 −/− and endothelial K IR 2.1 −/− mice using state-of-the-art live animal imaging where the local tissue O 2 environment was manipulated using a custom gas chamber. Second-by-second capillary RBC flow responses were recorded as O 2 was altered. A stepwise drop in PO 2 at the muscle surface increased RBC supply in capillaries of control animals while elevated O 2 elicited the opposite response; capillaries were confirmed to express Cx40. The RBC flow responses were rapid and tightly coupled to O 2 ; computer simulations did not support hypoxia as a driving factor. In contrast, RBC flow responses were significantly diminished in Cx40 −/− mice. Endothelial K IR 2.1 −/− mice, on the other hand, reacted normally to O 2 changes, even when the O 2 challenge was targeted to a smaller area of tissue with fewer capillaries. Conclusively, microvascular O 2 responses depend on coordinated electrical signaling via Cx40 gap junctions, and endothelial K IR 2.1 channels do not initiate the event. These findings reconceptualize the paradigm of blood flow regulation in skeletal muscle and how O 2 triggers this process in capillaries independent of extracellular K + .

Background Normal brain function depends on the ability of the vasculature to increase blood flow to regions with high metabolic demands. Impaired neurovascular coupling, such as the local hyperemic response to neuronal activity, may contribute to poor neurological outcome after stroke despite successful recanalization, that is, futile recanalization. Methods and Results Mice implanted with chronic cranial windows were trained for awake head‐fixation before experiments. One‐hour occlusion of the anterior middle cerebral artery branch was induced using single‐vessel photothrombosis. Cerebral perfusion and neurovascular coupling were assessed by optical coherence tomography and laser speckle contrast imaging. Capillaries and pericytes were studied in perfusion‐fixed tissue by labeling lectin and platelet‐derived growth factor receptor β. Arterial occlusion induced multiple spreading depolarizations over 1 hour associated with substantially reduced blood flow in the peri‐ischemic cortex. Approximately half of the capillaries in the peri‐ischemic area were no longer perfused at the 3‐ and 24‐hour follow‐up (45% [95% CI, 33%–58%] and 53% [95% CI, 39%–66%] reduction, respectively; P <0.0001), which was associated with contraction of an equivalent proportion of peri‐ischemic capillary pericytes. The capillaries in the peri‐ischemic cortex that remained perfused showed increased point prevalence of dynamic flow stalling (0.5% [95% CI, 0.2%–0.7%] at baseline, 5.1% [95% CI, 3.2%–6.5%] and 3.2% [95% CI, 1.1%–5.3%] at 3‐ and 24‐hour follow‐up, respectively; P =0.001). Whisker stimulation at the 3‐ and 24‐hour follow‐up led to reduced neurovascular coupling responses in the sensory cortex corresponding to the peri‐ischemic region compared with that observed at baseline. Conclusions Arterial occlusion led to contraction of capillary pericytes and capillary flow stalling in the peri‐ischemic cortex. Capillary dysfunction was associated with neurovascular uncoupling. Neurovascular coupling impairment associated with capillary dysfunction may be a mechanism that contributes to futile recanalization. Hence, the results from this study suggest a novel treatment target to improve neurological outcome after stroke.