Cardiocap 5

Heart rate was measured with a three‐lead electrocardiogram (ECG, Cardiocap/5; GE Healthcare, Waukesha, WI). Beat‐to‐beat MAP, SBP, and DBP were measured with a photoplethysmographic finger cuff (Finometer; FMS, Arnhem, Netherlands). A semi‐automated upper arm cuff (Dinamap; GE Medical System, Milwaukee, WI) was used to obtain three baseline blood pressure measurements, which were used to calibrate the baseline finger cuff signal in offline analysis. MAP, SBP, DBP, and HR were measured continuously at baseline and during handgrip.

All study protocols were conducted in the supine or left lateral position in a clinical research laboratory at 20–22°C. A three‐lead EKG (Cardiocap/5; GE Healthcare) to monitor HR was placed, as well as a finger BP cuff (Finometer, FMS), a pneumotrace to monitor respiratory movement, and an intravenous catheter in each arm. Prior to each stressor, three resting BPs were obtained by automated oscillometry of the right brachial artery (Philips Sure Signs VS3) after 15 min of quiet rest and these were used to verify the Finometer values as previously described (Muller et al. 2014b). All beat‐by‐beat variables were collected at 200 Hz by PowerLab (ADInstruments). CBVpeak and VTI in the distal left anterior descending coronary artery were obtained from the adjusted apical four‐chamber view using a GE Vivid 7 echocardiography system (all images acquired by Z. Gao). The specific procedures for measuring CBVpeak and in the LAD have been previously described by our laboratory (Momen et al. 2009; Ross et al. 2014). In brief, CBVpeak was calculated as the peak diastolic velocity (average of 3 or more cardiac cycles) and VTI was calculated as the area under the entire diastolic blood flow profile of the highest quality image for a single cardiac cycle. This approach is consistent with other published reports (Saraste et al. 2001; Meimoun et al. 2006; Abreu et al. 2014).

Individuals refrained from alcohol, caffeine, and exercise for 24 h prior to the study. After fasting for four hours, participants were admitted to the Clinical Research and Trials Unit at the Mayo Clinic. Participants rested quietly in the supine position during instrumentation and throughout the study. Baseline peripheral blood pressure was established using a brachial cuff. The average of three readings for peripheral blood pressure, each separated by two minutes, was recorded. Heart rate was recorded continuously using a 3‐lead ECG (Cardiocap/5; Datex‐Ohmeda, Louisville, CO).

The experiments were performed according to the NIH Guideline for the use of laboratory animals and were approved by the research ethics committee at the University of Groningen. Twelve male Wistar rats from Harlan weighing between 180–200 gr were randomly assigned to control or FTY720 treatment. Rats in the active treatment group received daily 1 mg/kg (maximal effect dose in rats[21 (link)]) body weight FTY720 (Novartis, Switzerland) via drinking water for 6 weeks. A week prior to treatment, animals were housed under standard conditions for acclimation at the animal facilities of the University of Groningen. During the study, ad libitum food and tap water were provided. After six weeks of treatment, rats were anesthetized with 2.5% isoflurane in oxygen and data of hemodynamic function recorded by a pressure transducer catheter, which was inserted into the carotid artery (Datex-Ohmeda, Cardiocap/5, USA). Afterwards, mesenteric arteries were isolated and transferred in normal physiologic saline for further studies.

On the morning of the study, participants awoke at 06.15 h and a retrograde intravenous catheter was placed in the dorsum of the non‐dominant hand under local anesthesia (2% lidocaine) for periodic blood sampling. Participants wore a mask connected to a two‐way non‐rebreathing valve for administration of gasses and to allow for continuous measurement of tidal volume, breathing frequency (Universal Ventilation Meter, Ventura, CA), and inspired/expired gasses (Cardiocap/5, Datex‐Ohmeda, Louisville, CO).

Systolic blood pressure (SBP), diastolic blood pressure (DBP), MAP, and HR were monitored continuously using a Datex-Ohmeda Cardiocap 5 (Datex-Ohmeda Inc, Madison, WI). Cardiac output (CO) was continuously monitored using a Vigilance II Monitor (Edwards Lifesciences, Irvine, CA). Measurements were documented manually every 5 min until the end of bleeding, and then every 20 min during the observation period.