Cerebral Blood Flow Response to Simulated Hypovolemia in Essential Hypertension

Supplemental Digital Content is available in the text.

Participants were screened by 12 lead ECG, a Siemens Multistix urine dipstick, clinic and ambulatory blood pressure monitoring, as well as height, weight, hip circumference and testing for orthostatic intolerance.
Participants were excluded if they were diagnosed with any cardiovascular (other than hypertension and hypertensive left ventricular hypertrophy), autonomic, respiratory, renal, cognitive or immunological disorders, BMI >35, or if they have a hip circumference >120cm due to the practical constraint of the lower body negative pressure chamber.

MR imaging parameters
Cerebral images were acquired with a 12 channel Head Matrix coil (Model 07577732, Siemens Heathineers), whilst cardiac images were acquired with an 8 channel Body Matrix coil (Model 07579555, Siemens Heathineers).

MR image analysis
The arteries were contoured in each frame of reconstructed flow from the smoothed magnitude images (see supplemental Figure S1). Mean flow velocity and total flow were quantified in each vessel using semi-automated Siemens software (Argus, Siemens Healthineers). Total CBF (tCBF) was calculated by summing the flow in the basilar and the internal carotid arteries. CBF was normalised to total brain tissue volume (nCBF) and presented as millilitres per 100g of tissue per minute. The brain tissue volumes were calculated from the T1-MPRAGE using FAST segmentation (FSL software; [1]) and the number of voxels for each tissue type quantified using FSLSTATS. The volumes were converted to grams assuming that 1cm 3 brain tissue weighs 1.036 grams [2].

About Lower Body Negative Pressure
Lower body negative pressure (LBNP) is a technique used to manipulate central blood volume, and has been validated as a model for haemorrhage in the baboon [3]. Typically, the lower body is encased in an airtight chamber from the level of the waist or iliac crest and a suction device used to manipulate the pressure within the chamber relative to athmospheric pressure.
LBNP is known to cause a large variety of physiological changes, including decreased central venous pressure, reduced preload of the heart, reduced stroke volume, reduced cardiac output and baro-reflex induced increases in heart rate, total peripheral resistance, muscle sympathetic nerve activity and the release of vasoactive and volume regulating endocrine factors, which are typically mimicking the effects of hypovolemia due to blood loss [4]. For example, Rickards et al. [5] showed that LBNP at -45mmHg and blood loss of up to 1000mL resulted in similar reductions in cerebral blood flow velocity and neither induced changes in MAP.
Hanson et al. [6] showed that the reduction in stroke distance at -20mmHg LBNP was equivalent to a blood loss of ~450mL. Finally, Hinjosa-Laborde et al. [3] suggested that LBNP at approximately -20mmHg equated to a blood loss of ~6.25% of the total blood volume, -40mmHg LBNP equated to a blood loss of ~12.5% of the total blood volume, and -50mmHg LBNP equated to a blood loss of ~18.75% of the total blood volume in the baboon model. Furthermore, Hinjosa-Laborde et al. [3]showed similar physiological reduction in pulse pressure, central venous pressure, systolic blood pressure, heart rate, stroke volume, cardiac output, vascular resistance, as well as similar changes in renin activity, blood bicarbonate, blood lactate, blood pH, partial pressure of oxygen, partial pressure of CO 2 , blood osmolarity, blood sodium levels, blood urea nitrogen, and glucose content between LBNP and blood loss.

Intra-and Inter-observer variability
Inter-observer variability of flow measurements from the ICA was performed by an independent and blinded researcher (SN and MDK) on 10 randomly selected participants. Intra-observer variability of the same ICA measurements was performed in 20 randomly selected participants. As a surrogate marker of vascular remodelling, arterial stenosis was assessed from the time of flight angiograms.

Methods
A trained radiologist (>8 years experience) assessed the time of flight angiograms evaluating the presence of internal carotid and vertebral artery stenosis. The data was reviewed in 3 orthogonal multiplanar-reformatted planes with cross-referencing of images. Maximum intensity projection images were generated and reviewed. Vessels were interrogated for the presence of focal atherosclerotic stenosis. Where present, a measurement of percentage area stenosis relative to adjacent normal calibre proximal and distal vessel was made. Table S1 with the rates of stenosis in each group.

Data by Sex
Given that there was no difference between the groups, all participants were pooled and grouped according to sex (male versus female). There was no difference between the sxes for tCBF during LBNP (p=0.28), MAP during LBNP (p=0.19), nor in the percentage of the CO diverted to the tCBF during LBNP (p=0.22). However, CO was affected differently in male and female volunteers (p=0.01, 11% of the total variation) and showed an interaction between LBNP and sex (p=0.01, 0.5% of the total variation). Post hoc CO was significantly lower in female volunteers at -40mmHg (p=0.045) and -50mmHg (p=0.0055) LBNP. This data is shown in S5 below.