BOLD fMRI and hemodynamic responses to somatosensory stimulation in anesthetized mice: spontaneous breathing vs. mechanical ventilation

Abstract

Mouse functional MRI (fMRI) has been of great interest due to the abundance of transgenic models. Due to a mouse's small size, spontaneous breathing has often been used. Because the vascular physiology affecting fMRI might not be controlled normally, its effects on functional responses were investigated with optical intrinsic signal (OIS) imaging and 9.4 T BOLD fMRI. Three conditions were tested in C57BL/6 mice: spontaneous breathing under ketamine and xylazine anesthesia (KX), mechanical ventilation under KX, and mechanical ventilation under isoflurane. Spontaneous breathing under KX induced an average pCO 2 of 83 mmHg, whereas a mechanical ventilation condition achieved a pCO 2 of 37-41 mmHg within a physiological range.The baseline diameter of arterial and venous vessels was only 7%-9% larger with spontaneous breathing than with mechanical ventilation under KX, but it was much smaller than that in normocapnic isoflurane-anesthetized mice. Three major functional studies were performed. First, CBV-weighted OIS and arterial dilations to 4-second forepaw stimulation were rapid and larger at normocapnia than hypercapnia under KX, but very small under isoflurane. Second, CBV-weighted OIS and arterial dilations by vasodilator acetazolamide were measured for investigating vascular reactivity and were larger in the normocapnic condition than in the hypercapnic condition under KX. Third, evoked OIS and BOLD fMRI responses in the contralateral mouse somatosensory cortex to 20-second forepaw stimulation were faster and larger in the mechanical ventilation than spontaneous breathing. BOLD fMRI peaked at the end of the 20-second stimulation under hypercapnic spontaneous breathing, and at 9 seconds under mechanical ventilation. The peak amplitude of BOLD fMRI was 2.2% at hypercapnia and~3.4% at normocapnia. Overall, spontaneous breathing induces sluggish reduced hemodynamic and fMRI responses, but it is still viable for KX anesthesia due to its simplicity, noninvasiveness, and well-localized BOLD activity in the somatosensory cortex.Abbreviations used: CBF, cerebral blood flow; CBV, cerebral blood volume; CBVw, CBV-weighted; EPI, echo planar imaging; GLM, general linear model; HR, heart rate; Iso, isoflurane; KX, ketamine and xylazine; MABP, mean arterial blood pressure; MV, mechanical ventilation; OIS, optical intrinsic signal; PCO 2 , partial arterial pressure of CO 2 ; PO 2 , partial arterial pressure of O 2 ; R, reflectance image intensity; ROI, region of interest; S1FL, primary somatosensory forelimb; SaO 2 , arterial oxygen saturation level; SB, spontaneous breathing. anesthesia, BOLD fMRI, cerebral blood volume, forepaw stimulation, isoflurane, ketamine, vascular reactivity | INTRODUCTIONBlood oxygenation level-dependent (BOLD) functional MRI (fMRI) in rodents has been used to understand the biophysics and neural sources of fMRI signals and to investigate brain functions. BOLD fMRI of anesthetized mice has been of particular interest due to the abundance of transgenic models, easy manipulation of genomes, and cost eff...

Characteristics of fMRI responses to visual stimulation in anesthetized vs. awake mice

“…Each mouse with trimmed whiskers was placed in a customized cradle with earplugs and a bite bar. To maintain anesthesia during the scan, supplementary doses (25 mg/kg ketamine and 1.25 mg/kg xylazine) were infused intermittently (every 30 or 45 minutes) through an IP line, as deemed necessary based on changes in physiological parameters (Jung et al, 2019; Shim et al, 2018; Shim et al, 2020).…”

“…However, the results of such studies have been inconsistent, and bilateral activity caused by cardiovascular responses has frequently been reported (Reimann et al, 2018) when only the contralateral hemisphere was expected to be active. Recently, we found that ketamine/xylazine anesthesia produces reliable localized somatosensory-induced fMRI responses (Jung, Shim, & Kim, 2019; Shim et al, 2018; Shim, Lee, & Kim, 2020) without changing the cardiac pulsations or respiration rate (Shim et al, 2018). Thus, we used ketamine/xylazine anesthesia for this mouse fMRI study to characterize the BOLD fMRI responses to visual stimulation.…”

Systematic Review: Anaesthetic Protocols and Management as Confounders in Rodent Blood Oxygen Level Dependent Functional Magnetic Resonance Imaging (BOLD fMRI)–Part A: Effects of Changes in Physiological Parameters

“…A recent study (published after the date the systematic search was performed) in mechanically ventilated vs. spontaneously breathing mice measured vessel diameter with optic intrinsic signal imaging as well as BOLD response to electrical paw stimulation. At the average p a CO 2 of 83 mmHg in spontaneously breathing animals, baseline vessel diameter was significantly larger, and increase in vessel diameter as well as the peak BOLD response to paw stimulation were significantly reduced and delayed compared to values measured at an average p a CO 2 of 41 in mechanically ventilated animals (Shim et al, 2020). One of the included interventional studies however observed a significant decrease of response to stimulation already with 1% CO 2 admixture, corresponding to an increase in p a CO 2 of 5 mmHg (Nasrallah et al, 2015).…”

“…While both routes may be used in terminal studies, the less invasive oropharyngeal route should be chosen for recovery studies. While rats appear to recover with few complications from oropharyngeal intubation (Rivard et al, 2006), a recent publication described cessation of weight gain in the first week after extubation in mice (Shim et al, 2020). One of the authors (AS) has however successfully recovered mice after intubation and mechanical ventilation, suggesting that longitudinal studies with appropriate ventilatory management are feasible in mice, although the technical challenges may require an experienced operator.…”

Systematic Review: Anesthetic Protocols and Management as Confounders in Rodent Blood Oxygen Level Dependent Functional Magnetic Resonance Imaging (BOLD fMRI)—Part B: Effects of Anesthetic Agents, Doses and Timing

“…However, isoflurane may not be the ideal agent. It has strong vasodilatory effects resulting in slower and weaker hemodynamic responses to stimulation [ 144 ]. Moreover, unspecific bilateral BOLD signal changes in unilateral paw stimulation studies in mice have been reported [ 38 , 122 ], as well as pronounced differences of fc patterns under isoflurane to those found in awake rats [ 121 ].…”