Two‐photon microscopy enables non‐invasive, in vivo, longitudinal study of arterial Ca 2+ and diameter in optical biosensor mice during the development of hypertension (856.7)

Here, we report that use of two‐photon microscopy of ear arteries in exMLCK biosensor mice enables non‐invasive, in vivo, serial examination of vascular smooth muscle Ca 2+ during the development of hypertension. ExMLCK mice express a FRET‐type biosensor of Ca 2+ /Calmodulin based on smooth muscle myosin light chain kinase (MLCK) and its ratiometric properties permit direct quantification of intracellular Ca 2+ concentration. Anesthetized biosensor mice were positioned on a custom platform, facilitating imaging of selected arterial branches within the intact depilated ear, and studied before and during chronic infusion of angiotensin‐II (ANGII; 800 µg/g/day) via mini‐osmotic pump. Acute responsiveness to i.p. phenylephrine (PE; 30µg/g) was determined in a separate group of mice equipped with arterial blood pressure radio telemetry. Eight days of chronic ANGII increased intracellular Ca 2+ from 294±47nM to 501±92nM ( P <0.05) and decreased artery diameter by ‐14±2% ( P <0.05). Acute PE induced robust vasomotion and 2‐3 fold oscillations in spatially averaged Ca 2+ . In summary, this new methodology permits for the first time the simultaneous study of Ca 2+ signaling, structural remodeling and vascular reactivity in the intact living mouse during hypertension development. Grant Funding Source : Supported by NHLBI RO1091969, RO1HL107654, RO1DK077140, and T32HL072751.