Dual-color miniscope imaging of microvessels and neuronal activity in the hippocampus CA1 region of freely moving mice following alcohol administration.

Moderate-to-heavy episodic ("binge") drinking is the most common form of alcohol consumption in the United States. Alcohol at binge drinking concentrations reduces brain artery diameter in vivo and in vitro in many species including rats, mice, and humans. Despite the critical role played by brain vessels in maintaining neuronal function, there is a shortage of methodologies to simultaneously assess neuron and blood vessel function in deep brain regions. Here, we investigate cerebrovascular responses to ethanol by choosing a deep brain region that is implicated in alcohol disruption of brain function, the hippocampal CA1, and describe the process for obtaining simultaneous imaging of pyramidal neuron activity and diameter of nearby microvessels in freely moving mice via a dual-color miniscope. Recordings of neurovascular events were performed upon intraperitoneal injection of saline versus 3 g/kg ethanol in the same mouse. In male mice, ethanol mildly increased the amplitude of calcium signals while robustly decreasing their frequency. Simultaneously, ethanol decreased microvessel diameter. In females, ethanol did not change the amplitude or frequency of calcium signals from CA1 neurons but decreased microvessel diameter. A linear regression of ethanol-induced reduction in number of active neurons and microvessel constriction revealed a positive correlation (R = 0.981) in females. Together, these data demonstrate the feasibility of simultaneously evaluating neuronal and vascular components of alcohol actions in a deep brain area in freely moving mice, as well as the sexual dimorphism of hippocampal neurovascular responses to alcohol.

Differential Functional Contribution of BK Channel Subunits to Aldosterone-Induced Channel Activation in Vascular Smooth Muscle and Eventual Cerebral Artery Dilation.

Calcium/voltage-activated potassium channels (BK) control smooth muscle (SM) tone and cerebral artery diameter. They include channel-forming α and regulatory ß1 subunits, the latter being highly expressed in SM. Both subunits participate in steroid-induced modification of BK activity: ß1 provides recognition for estradiol and cholanes, resulting in BK potentiation, whereas α suffices for BK inhibition by cholesterol or pregnenolone. Aldosterone can modify cerebral artery function independently of its effects outside the brain, yet BK involvement in aldosterone's cerebrovascular action and identification of channel subunits, possibly involved in steroid action, remains uninvestigated. Using microscale thermophoresis, we demonstrated that each subunit type presents two recognition sites for aldosterone: at 0.3 and ≥10 µM for α and at 0.3-1 µM and ≥100 µM for ß1. Next, we probed aldosterone on SM BK activity and diameter of middle cerebral artery (MCA) isolated from ß1-/- vs. wt mice. Data showed that ß1 leftward-shifted aldosterone-induced BK activation, rendering EC50~3 µM and ECMAX ≥ 10 µM, at which BK activity increased by 20%. At similar concentrations, aldosterone mildly yet significantly dilated MCA independently of circulating and endothelial factors. Lastly, aldosterone-induced MCA dilation was lost in ß1-/- mice. Therefore, ß1 enables BK activation and MCA dilation by low µM aldosterone.