Mice were anesthetized by intraperitoneal injection of ketamine (80 mg/kg) and xylazine (10 mg/kg). A middle line incision was made at the neck, and the carotid artery was separated carefully. Then, a monofilament suture (Doccol, Sharon, MA, USA) was inserted into the internal carotid artery (ICA). Rectal temperature was maintained at 37.0°C during and after surgery with a temperature controlled heating pad. The regional cerebral blood flow (rCBF) was monitored using laser Doppler flowmetry (moor VMS-LDF2, Moor Instruments, Axminster, UK). Animals whose blood flow did not reduce to 20% were excluded. The monofilament suture was removed 1 hour for reperfusion.
Moorvms ldf2
Manufactured by Moor Instruments
Sourced in United Kingdom
The MoorVMS-LDF2 is a laser Doppler flowmeter that measures blood perfusion in tissue. It provides quantitative measurements of microvascular blood flow and tissue perfusion.
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19 protocols using moorvms ldf2
1
Transient Focal Cerebral Ischemia in Mice
2
Transient Middle Cerebral Artery Occlusion in Mice
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All the experimental protocols were approved by the Animal Care and Use Committee of the Shanghai Jiao Tong University School of Medicine, Shanghai, China. First, 12–14-week-old male C57BL/6 mice (Jiesijie, Shanghai, China) were anesthetized with 1.5–2% isoflurane, and rectal temperatures and cerebral blood flow (CBF) were monitored. Transient middle cerebral artery occlusion (tMCAO) was induced using intraluminal suture as described before (Zuo et al., 2019 (link)). A midline neck incision was made, and the left common carotid artery (CCA), external carotid artery (ECA), and internal carotid artery (ICA) were isolated. The ECA was ligated with 5-0 silk suture, and a 6-0 nylon monofilament coated with silicon resin (6023PK, Doccol Corp., Redlands, CA, USA) was inserted from ECA to occlude the MCA. The reduction of the CBF was confirmed by a laser-Doppler flowmeter (moorVMS-LDF-2, Moor Instruments, Devon, UK). Only animals with CBF lower than 20% of baseline were admitted in this study. One hour after occlusion, the suture was gently removed to allow reperfusion. Mice in the Sham group received the same surgical procedure without filament insertion. Body temperature was regularly monitored and maintained at 37 ± 0.5°C with a heating pad. All animals were allowed ad libitum access to water and food after surgery.
3
Transient Middle Cerebral Artery Occlusion in Mice
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Ischemic stroke was induced in female C57BL/6JRj mice by a transient middle cerebral artery occlusion (tMCAo), which is mimicking one of the most common types of ischemic stroke in patients (Engel et al., 2011 (link)). The intraluminal occlusion model was performed as described elsewhere with minor modifications (Engel et al., 2011 (link); Zinnhardt et al., 2015 (link)). In short, a 7–0 monofilament (tip diameter 190–200 ml, coating length 2–3 mm, 70SPRePK5, Doccol Corp., Sharon, MA, United States) was inserted in the right common carotid artery and placed to block blood supply via the middle cerebral artery. The filament was held in place for 30 min followed by retraction of the filament leading to reperfusion. A Laser Doppler probe (moorVMS-LDF2, Moor Instruments, United Kingdom) was placed on the skull of the mice to monitor cerebral blood flow, considering a drop of ≥ 80% CBF as a successful stroke induction. Animals were anesthetized during the whole time of surgery, using 1.5% isoflurane (Abbott Animal Health, AbbottPark, IL, United States) in a 2:1 air and oxygen mixture.
4
Multimodal Physiological Monitoring
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Similar to the CPT study, the measurement setup involved the acquisition of PPG signals from the ear canal and the left index finger, ECG signal, and temperature measurements from the periphery (skin) and the core (tympanic). PPG and ECG measurements were made using the ZenPPG system. Core temperature was measured using a tympanic thermometer (ThermoScan-5 IRT4520, Braun GmbH, Frankfurt, Germany). Skin temperature was measured using the laser Doppler flowmeter (LDF) (moorVMS-LDF2, Moor Instruments, Devon, UK).
5
Cerebral Blood Flow Monitoring
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To ensure that the ischemic insult was uniform among all experimental animals, cerebral blood flow was measured with a laser Doppler perfusion and temperature monitor (moorVMS-LDF2, Moor Instruments, Devon, UK) immediately after the filament was advanced into the middle cerebral artery. Briefly, the skull was exposed, and whole brain scans were conducted using the blood flow monitor in each cerebral hemisphere. Digital images were consecutively acquired, and the average instantaneous blood flow was calculated using LDF2 software (Moor Instruments). Cerebral blood flow decreased to 30–35% and remained relatively stable (Figure 2
6
Laser Doppler Measurement of Skin Blood Flow
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Skin blood flow (SBF) was determined via a Laser Doppler and Perfusion Monitor (moorVMS-LDF2, Moor Instruments, Axminster, UK) with the VP1/7 probe placed on the anterior aspect of rectus femoris muscle over the middle of the muscle belly. Laser Doppler flowmetry is based on red blood cell flux and calculated as product of red blood cell concentration and velocity. This variable is expressed in arbitrary units (PU) and is indicative of superficial skin blood flow [18 ].
7
Transient Global Ischemia Rat Model
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Transient global ischemia was induced by using the 4-VO method with minor modifications (Pulsinelli et al., 1982 (link); Han et al., 2016 (link)). Briefly, rats were anesthetized by intraperitoneal (i.p.) injection of 400 mg/kg chloral hydrate (Fluka, Buchs, Switzerland). Body temperature was maintained at 37.0 ± 0.2°C during surgery using heating pads. The bilateral vertebral arteries were exposed and permanently electrocauterized. Both common carotid arteries (CCAs) were occluded with carotid artery clips for 15 min, and blood flow was monitored at a point in the skull (bregma: −2.0 mm, midline: 5.0 mm) with a probe connecting a laser-Doppler flowmetry device (MoorVMS-LDF2, Moor Instruments, Devon, UK) before, during and after ischemia. The ischemic model was considered successful when the cerebral blood flow decreased to 10%–15% of pre-ischemic levels and returned to the baseline (100 ± 10%) after release of the carotid artery clips. Sham animals underwent all aspects of handling and surgery without occlusion of the vertebral and CCAs.
8
Measurement of Renal Blood Flow
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In this set of experiments, the animals were anaesthetized with ketamine/xylazine (90/15 mg/kg, i.m.) and a transverse abdominal incision was performed to assess the posterior right subhepatic space, allowing for the visualization of the right kidney. A laser probe (model VP1) connected to a laser Doppler blood flow monitor (moorVMS-LDF2, Moor Instruments, England) was carefully placed directly onto the kidney, allowing for the measurement of renal blood flow (in arbitrary units). The probe was kept in this position, and the surgical incision was covered with gauze sponges soaked in sterile phosphate-buffered saline to protect the kidney from drying out. The laser probe remained tightly fixed on the kidney surface, and very stable and constant trace recordings of renal blood flow were obtained. During the experiments, the animals were maintained on a warming pad and allowed to breathe spontaneously. An interval of 20 min was observed before the measurement of basal values and renal blood flow was measured for an additional 15 min [18 (link)].
9
Transient Focal Cerebral Ischemia Model
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Following the methodology described in Longa et al. (1989 (link)), under microscopy, blunt dissection was performed through the carotid triangle to expose the left common carotid artery (CCA) and bifurcation of the external carotid artery (ECA) and internal carotid artery (ICA). The superior thyroid artery (STA) of the ECA was then isolated and coagulated, followed by the occipital artery. The ICA was carefully isolated from the adjacent vagus nerve. Next, two 3–0 silk sutures were placed around the ECA with one ligated distal to the STA. Cerebral blood flow (CBF) was monitored and recorded for baseline readings with the LDF system (moorVMS-LDF2, Moor Instruments) for 5 min. Two micro-vascular clips were placed on the CCA and ICA to prevent any bleeding, and the ECA was partially severed with microscissors. A 4–0 monofilament nylon suture with an occlusive diameter of 0.41 mm was inserted through the proximal ECA into the ICA, and the remaining ECA was severed. The micro-vascular clip on the ICA was removed and the filament was advanced until the LDF showed a 70% decrease from baseline CBF. The silk suture around the ECA stump was tightened around the intraluminal nylon suture to prevent bleeding. Occlusion time was set for 90 min then the filament was removed to allow for reperfusion.
10
Laser Doppler Measurement of Skin Blood Flow
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