Laser speckle flowmetry method for measuring spatial and temporal hemodynamic alterations throughout large microvascular networks.

1) To develop and validate laser speckle flowmetry (LSF) as a quantitative tool for individual microvessel hemodynamics in large networks. 2) To use LSF to determine if structural differences in the dorsal skinfold microcirculation (DSFWC) of C57BL/6 and BALB/c mice impart differential network hemodynamic responses to occlusion. We compared LSF velocity measurements with known/measured velocities in vitro using capillary tube tissue phantoms and in vivo using mouse DSFWCs and cremaster muscles. Hemodynamic changes induced by feed arteriole occlusion were measured using LSF in DSFWCs implanted on C57BL/6 and BALB/c mice. In vitro, we found that the normalized speckle intensity (NSI) versus velocity linear relationship (R(2) ≥ 0.97) did not vary with diameter or hematocrit and can be shifted to meet an expected operating range. In vivo, DSFWC and cremaster muscle preparations (R(2) = 0.92 and 0.95, respectively) demonstrated similar linear relationships between NSI and centerline velocity. Stratification of arterioles into predicted collateral pathways revealed significant differences between C57BL/6 and BALB/c strains in response to feed arteriole occlusion. These data demonstrate the applicability of LSF to intravital microscopy microcirculation preparations for determining both relative and absolute hemodynamics on a network-wide scale while maintaining the resolution of individual microvessels.