Figure 1.
Time-series fluorescence imaging of indocyanine green (ICG) on a mouse head and generation of a cerebral blood flow (CBF) map.
(A) Schematic of imaging setup. (B) Time-series acquisition of fluorescence images after intravenous bolus injection of ICG. The scalp was excised and the bregma was positioned at the center of the images (scale bar: 2 mm). (C) ICG dynamics in three pixels after injection of an ICG bolus, indicated in (B). Different regions of interest are indicated by colored shapes (red circle, cerebrum; green rectangle, large vein; blue triangle, skeletal muscle). (D) Smoothed and interpolated dynamics were plotted as a solid line, and the arrival time (Tarrival) and the first peak time (Tpeak) for calculating the rising time (Trising) are indicated on the time axis. (E) A representative CBF map drawn using the Trising parameter. CCD, charge-coupled device; LED, light-emitting diode; a.u., arbitrary unit; Tmax, maximum intensity time.
Figure 2.
Intracranial blood flow parameters are predominantly measured by dynamic fluorescence imaging analysis.
(A) Image of a mouse head (in which the extracerebral tissues were totally removed in the left hemisphere) and a Trising map. To remove extracerebral tissues, the scalp was excised, and a craniotomy was applied along with dura mater excision. (B) ICG dynamics of a region from which all extracerebral tissues were removed (red line) and a contralateral region with intact skull and dura mater (blue line). (C and D) Representative Trising maps of a head with an intact scalp in an ICR mouse (C) and a C57Bl/6 mouse (D). (E and F) Diagrams of the selective block of blood supply to intracerebral (E) and extracerebral (F) tissues. (G and H) Left internal carotid artery (ICA) was ligated, and a nylon suture filament was inserted toward the middle cerebral artery (MCA) to block the intracerebral blood supply. The external carotid artery (ECA) was ligated to block the extracerebral blood supply. Trising maps for the selective block of the intracerebral blood supply in a scalp-removed mouse (G) and an intact-scalp mouse (H). (I and J) Trising maps for the selective block of the extracerebral blood supply in a scalp-removed mouse (I) and an intact-scalp mouse (J). All mice were anesthetized with ketamine and xylazine. CCA, common carotid artery.
Figure 3.
Types of CBF maps and a comparison of CBF parameters in normal and ischemic hemispheres.
(A) Representative CBF maps using the four parameters described in Table 1 were generated from a normal condition (left panels) and an ischemic condition in which the left MCA was occluded (right panels). (B) Average blood flow parameters of regions over left somatosensory cortices of nine mice before and after MCA occlusion (MCAO) surgery. Trising and blood flow index (BFI) parameters showed significant differences between the normal and ischemic conditions (t-test, ***p<0.001). (C) Average blood flow parameters of regions over ipsilateral ischemic somatosensory cortices were compared to those regions over contralateral normal cortices in nine mice during MCAO. All four parameters showed significant differences (paired t-test, ***p<0.001). MTT, mean transit time; Contra, contralateral region; Ipsi, Ipsilateral region.
Table 1.
Definitions and interpretations of cerebral blood flow (CBF) map parameters.
Figure 4.
Detection of cerebral hemodynamic changes using CBF maps.
(A) Representative Trising maps for mice anesthetized with either 1.5% isoflurane or 0.1 mg/g ketamine and 0.01 mg/g xylazine. (B) Averaged CBF parameters of regions over left somatosensory cortices of six mice anesthetized with isoflurane and six mice anesthetized with ketamine and xylazine. Tarrival, Trising, and MTT parameters increased significantly, and BFI parameter was decreased significantly, in ketamine and xylazine group (t-test, **p<0.01; ***p<0.001). (C) Timeline of the transient MCAO protocol (upper diagram) and representative Trising maps (lower panels). Reperfusion (RP) was induced at 45 min after MCA occlusion. The six time points for imaging acquisition are indicated under the timeline.