Cerebral blood flow and BOLD fMRI responses to hypoxia in awake and anesthetized rats

doi:10.1016/j.brainres.2006.11.097

Brain Research

Volume 1135, 2 March 2007, Pages 186-194

https://doi.org/10.1016/j.brainres.2006.11.097 Get rights and content

Abstract

This study investigated the functional MRI responses to graded hypoxia in awake/restrained and anesthetized animals by measuring cerebral blood flow (CBF) and blood oxygenation (BOLD) changes and estimating changes in cerebral metabolic rate of oxygen (CMRO₂). Hypoxia in isoflurane anesthetized rats reduced blood pressure but did not change heart rate and respiration rate. In contrast, hypoxia in awake animals showed compensatory responses by sustaining blood pressure, increasing heart rate and respiration rate. Basal CBF was higher under isoflurane anesthesia than awake state because isoflurane is a vasodilator. Graded hypoxia decreased BOLD signals. Surprisingly, hypoxia also decreased CBF likely because hypoxia induced hypocapnia. Hypoxia-induced CBF and BOLD decreases were smaller in awake, relative to anesthetized, rats at low pO₂, but similar at high pO₂. CBF leveled off with decreasing hypoxia-induced pCO₂ in awake rats, but monotonically decreased in anesthetized rats. CMRO₂ estimated using a biophysical BOLD model did not change under mild hypoxia but was reduced under severe hypoxia relative to baseline. These results showed that isoflurane attenuated automomic responses to hypoxia, hypoxia-induced hypocapnia dominated CBF changes, tissues in awake conditions appeared better oxygenated, and severe hypoxia reduced oxygen metabolism. This study underscored the marked differences in BOLD and CBF MRI responses to hypoxia in vivo between awake and anesthetized conditions and has implications for functional MRI studies of hypoxia in anesthetized animal models.

Introduction

The mammalian brain depends on a continuous and adequate supply of oxygen to maintain its structural and functional integrity. If oxygen delivery is sufficiently compromised, loss of consciousness could occur within seconds, irreversible neuronal damage and severe brain dysfunction, within minutes (Siesjo, 1978). During hypoxia, there are systematic autonomic cerebrovascular responses, such as vasodilation and increases in respiration rate, heart rate, cerebral blood volume, and cerebral blood flow (CBF), to compensate for reduced inspired oxygen. These autonomic responses depend on the severity and duration of hypoxia, species, tissue type, and level and type of anesthetic (Siesjo, 1978).

Combined CBF and blood-oxygenation-level-dependent (BOLD; Ogawa et al., 1990) functional MRI provides a unique means to investigate the neurovascular coupling under graded hypoxia non-invasively. Stimulus-evoked CMRO₂ changes can be estimated with combined BOLD and CBF measurements (Davis et al., 1998, Liu et al., 2004). While general anesthesia with mechanical ventilation is commonly used for immobilization in MRI studies of animal models, there has been increasing interest in imaging awake animals because of the important applications of awake models for mapping higher-order cognitive brain functions. The feasibility of performing fMRI under awake and restrained conditions has been demonstrated using the BOLD technique (Logothetis et al., 1999, Wyrwicz et al., 2000), MION (monocrystalline iron oxide nanoparticules) technique to detect stimulus-evoked changes in blood volume (Dubowitz et al., 2001, Vanduffel et al., 2001), and perfusion and perfusion-based functional MRI (Sicard et al., 2003). While fMRI studies of awake animals are generally challenging, they also offer many distinct advantages. First, the confounding effects of anesthesia on blood flow, metabolism and neuro-vascular coupling can be avoided. Second, general neural activity is not suppressed, which leads to more salient fMRI signal changes. Finally, sub-cortical and higher order cognitive functions can be studied in the awake model. Such studies would be very difficult if not impossible under anesthesia. The disadvantages of performing fMRI studies on awake animals include motion artifacts and restraint stress. Characterizing the differences and similarities in physiological and functional measures between awake and anesthetized conditions could help to better understand the fMRI signals and improve study designs.

The goal of this study was to investigate the cerebrovascular responses to graded hypoxia in awake and anesthetized (2% isoflurane) spontaneously breathing rats. CBF and BOLD fMRI signals were simultaneously measured using the continuous arterial spin-labeling technique with echo-planar-imaging acquisition. Cerebral metabolic rate of oxygen (CMRO₂) was estimated using the biophysical BOLD model. MRI data were correlated with blood pressure, heart rate, respiration rate, and blood gases in the same animals.

Section snippets

Physiological parameters

MABP, heart rate, and respiration rate under basal conditions and graded hypoxia are plotted in Fig. 1. Under awake basal (21% O₂) conditions, the respiration rate, heart rate, and MABP were within normal physiological ranges (Sharp and LaRegina, 1998), consistent with previously reported values (Sicard et al., 2003). These parameters under anesthetized basal condition were clearly reduced (P < 0.05) but remained within normal physiological ranges (Sharp and LaRegina, 1998). During graded hypoxia

Discussion

Relative to normoxia, hypoxia (i.e., 9% O₂) in 2%-isoflurane anesthetized rats reduces in blood pressure and does not change heart and respiration rate. In contrast, hypoxia in awake animals shows compensatory responses by sustaining blood pressure, and increasing heart rate and respiration rate. Hypoxia induces hypocapnia. Basal CBF under anesthesia is higher than under the awake state because isoflurane is a potent vasodilator. Graded hypoxia decreases BOLD signals. Surprisingly, hypoxia

Animal preparation

These studies were approved and closely monitored by Institutional Animal Care and Use Committee (IACUC). Seven Sprague–Daley rats (250–300 g) were imaged under awake and anesthetized (2% isoflurane) conditions in the same animals. Rats were initially anesthetized with 2% isoflurane. A femoral artery was catheterized for monitoring the mean arterial blood pressure (MABP), heart rate (HR), and for sampling blood gases. Topical anesthetics (2% lidocaine) was applied to the surgical site which was

Acknowledgments

This work was supported in part by the NIH (NINDS, R01-NS45879), the American Heart Association (SDG-0430020N), and the Whitaker Foundation (RG-02-0005).

References (35)

J.A. King et al.
Procedures for minimizing stress for fMRI studies in conscious rats
J. Neurosci. Methods
(2005)
R.P. Shockley et al.
Determination of rat cortical blood volume changes by capillary mean transit time analysis during hypoxia, hypercapnia and hyperventilation
Brain Res.
(1988)
K.M. Sicard et al.
Effects of hypoxia, hyperoxia and hypercapnia on baseline and stimulus-evoked BOLD, CBF and CMRO2 in spontaneously breathing animals
NeuroImage
(2005)
W. Vanduffel et al.
Visual motion processing investigated using contrast agent-enhanced fMRI in awake behaving monkeys
Neuron.
(2001)
G. Bao et al.
Acute blood pressure elevation during repetitive hypocapnic and eucapnic hypoxia in rats
J. Appl. Physiol.
(1997)
D. Bereczki et al.
Hypoxia increases velocity of blood flow through parenchymal microvascular systems in rat brain
J. Cereb. Blood Flow Metab.
(1993)
J.L. Boxerman et al.
MR contrast due to intravascular magnetic susceptibility perturbations
Magn. Reson. Med.
(1995)
T.L. Davis et al.
Calibrated functional MRI: mapping the dynamics of oxidative metabolism
Proc. Natl. Acad. Sci. U. S. A.
(1998)
D.J. Dubowitz et al.
Enhancing fMRI contrast in awake-behaving primates using intravascular magnetite dextran nanoparticles
NeuroReport
(2001)
T.Q. Duong et al.
Functional MRI of calcium-dependent synaptic activity: cross correlation with CBF and BOLD measurements
Magn. Reson. Med.
(2000)

T.Q. Duong et al.

Effect of hyperoxia, hypercapnia and hypoxia on cerebral interstitial oxygen tension and cerebral blood flow in the rat brain: an 19F/1H study

Magn. Reson. Med.

(2001)

Q. Duong et al.

Localized blood flow response at sub-millimeter columnar resolution

Proc. Natl. Acad. Sci. U. S. A.

(2001)

J.O. Eichling et al.

Evidence of the limitations of water as a freely diffusible tracer in brain of the Rhesus monkey

Circ. Res.

(1974)

R.L. Grubb et al.

The effects of changes in PaCO2 on cerebral blood volume, blood flow, and vascular mean transit time

Stroke

(1974)

P. Herscovitch et al.

What is the correct value for the brain–blood partition coefficient for water?

J. Cereb. Blood Flow Metab.

(1985)

W.E. Hoffman et al.

Cerebral autoregulation in awake versus isoflurane-anesthetized rats

Anesth. Analg.

(1991)

R.D. Hoge et al.

Linear coupling between cerebral blood flow and oxygen consumption in activated human cortex

Proc. Natl. Acad. Sci.

(1999)

Cited by (68)

Interindividual Variability of Functional Connectivity in Awake and Anesthetized Rhesus Macaque Monkeys
2019, Biological Psychiatry: Cognitive Neuroscience and Neuroimaging
Nonhuman primate (NHP) models are commonly used to advance our understanding of brain function and organization. However, to date, they have offered few insights into individual differences among NHPs. In large part, this is due to the logistical challenges of NHP research, which limit most studies to 5 subjects or fewer.
We leveraged the availability of a large-scale open NHP imaging resource to provide an initial examination of individual differences in the functional organization of the NHP brain. Specifically, we selected one awake functional magnetic resonance imaging dataset (Newcastle University: n = 10) and two anesthetized functional magnetic resonance imaging datasets (Oxford University: n = 19; University of California, Davis: n = 19) to examine individual differences in functional connectivity characteristics across the cortex as well as potential state dependencies.
We noted significant individual variations of functional connectivity across the macaque cortex. Similar to the findings in humans, during the awake state, the primary sensory and motor cortices showed lower variability than the high-order association regions. This variability pattern was significantly correlated with T1-weighted and T2-weighted mapping and degree of long-distance connectivity, but not short-distance connectivity. The interindividual variability under anesthesia exhibited a very distinct pattern, with lower variability in medial frontal cortex, precuneus, and somatomotor regions and higher variability in the lateral ventral frontal and insular cortices.
This work has implications for our understanding of the evolutionary origins of individual variation in the human brain and methodological implications that must be considered in any pursuit to study individual variation in NHP models.
Hemodynamic and neuronal responses to cocaine differ in awake versus anesthetized animals: Optical brain imaging study
2019, NeuroImage
Citation Excerpt :
The observed dilation was larger in arteries than in veins likely due to more muscular structures in their vessel walls. As expected from the observed vasodilation, isoflurane also increased CBFv (Timothy, 2007). Isoflurane also depressed Ca2+ signaling, consistent with inhibition of neuronal activity during isoflurane's anesthesia (Wu et al., 2016), which is also in agreement with prior studies showing that isoflurane inhibited neuronal activities (Baumgart et al., 2015, Study, 1994).
Cocaine is a highly addictive drug with complex pharmacological effects. Most preclinical imaging studies investigating the effects of cocaine in the brain have been performed under anesthesia, which confounds findings. To tackle this problem, we used optical imaging to compare the effects of cocaine in the awake versus the anesthetized states. For this purpose, we customized an air floating mobile cage to fit the multi-wavelength spectral and laser speckle optical imaging system and implanted a multi-layer cranial window over the mouse somatosensory cortex. Results showed significant differences in neuronal activity and hemodynamics at baseline and in response to cocaine between the awake and the anesthetized states (isoflurane anesthesia). Specifically, 1) at baseline isoflurane dilated cerebral vessels, increased cerebral blood flow and depressed neuronal Ca²⁺ activity compared to the awake state; 2) acute cocaine (1 mg/kg iv) vasoconstricted blood vessels (arteries and veins) and decreased cerebral blood flow and oxygenated hemoglobin in the anesthetized state but not in the awake condition; 3) cocaine increased the accumulation of mean intracellular Ca²⁺ in neurons in the anesthetized state but not in the awake condition; and 4) in the awake state acute cocaine increased neuronal activities (increased the frequency of Ca²⁺ transients) and increased neuronal synchronization. We also corroborated that in the awake state cocaine also disrupted neurovascular coupling. These findings indicate that both vascular and neuronal responses to cocaine are influenced by isoflurane anesthesia, which highlights the importance of imaging awake animals when studying the effects of cocaine or other drugs in the brain.
Bacterial lipopolysaccharide-induced systemic inflammation alters perfusion of white matter-rich regions without altering flow in brain-irrigating arteries: Relationship to blood-brain barrier breakdown?
2018, Journal of Neuroimmunology
Citation Excerpt :
Therefore, and even though signs of vasospasms have been observed with TOF MRA in septic patients with encephalopathy (Bartynski et al., 2006; Polito et al., 2013), progressively increasing circulating concentrations of bacterial LPS after its ip injection did not alter flow in the dorsal forebrain-irrigating arteries of rodents. The increase in blood perfusion of the cortex and striatum with time, but independently from treatment, as found in the present work, is in accordance with previous studies employing ASL in intact rats under isoflurane anesthesia (Sicard et al., 2003; Duong, 2007). This isoflurane-induced hyperemia is due to increased cerebral NO production (Kehl et al., 2002, Stejskal and Tanner, 1965) and even more important for other inhaled anesthetics (Holmstrom and Akeson, 2005).
To better understand brain dysfunction during sepsis, cerebral arterial blood flow was assessed with Phase Contrast Magnetic Resonance Imaging, perfusion with Arterial Spin Labeling and structure with diffusion-weighted Magnetic Resonance Imaging in rats after intraperitoneal administration of bacterial lipopolysaccharides. Although cerebral arterial flow was not altered, perfusion of the corpus callosum region and diffusion parallel to its fibers were higher after lipopolysaccharide administration as compared to saline injection. In parallel, lipopolysaccharide induced perivascular immunoglobulin-immunoreactivity in white matter. These findings indicate that systemic inflammation can result in increased perfusion, blood-brain barrier breakdown and altered water diffusion in white matter.
The multi-level impact of chronic intermittent hypoxia on central auditory processing
2017, NeuroImage
Citation Excerpt :
In this study, fMRI is performed in a normal atmosphere after CIH treatment. Therefore, the impact on fMRI signals is expected to be different from that due to acute or transient hypoxia (Duong, 2007; Sicard and Duong, 2005). No qualitative differences are observed in subject behavior or food intake during the CIH treatment.
During hypoxia, the tissues do not obtain adequate oxygen. Chronic hypoxia can lead to many health problems. A relatively common cause of chronic hypoxia is sleep apnea. Sleep apnea is a sleep breathing disorder that affects 3–7% of the population. During sleep, the patient's breathing starts and stops. This can lead to hypertension, attention deficits, and hearing disorders. In this study, we apply an established chronic intermittent hypoxemia (CIH) model of sleep apnea to study its impact on auditory processing. Adult rats were reared for seven days during sleeping hours in a gas chamber with oxygen level cycled between 10% and 21% (normal atmosphere) every 90 s. During awake hours, the subjects were housed in standard conditions with normal atmosphere. CIH treatment significantly reduces arterial oxygen partial pressure and oxygen saturation during sleeping hours (relative to controls). After treatment, subjects underwent functional magnetic resonance imaging (fMRI) with broadband sound stimulation. Responses are observed in major auditory centers in all subjects, including the auditory cortex (AC) and auditory midbrain. fMRI signals from the AC are statistically significantly increased after CIH by 0.13% in the contralateral hemisphere and 0.10% in the ipsilateral hemisphere. In contrast, signals from the lateral lemniscus of the midbrain are significantly reduced by 0.39%. Signals from the neighboring inferior colliculus of the midbrain are relatively unaffected. Chronic hypoxia affects multiple levels of the auditory system and these changes are likely related to hearing disorders associated with sleep apnea.
Circulating bacterial lipopolysaccharide-induced inflammation reduces flow in brain-irrigating arteries independently from cerebrovascular prostaglandin production
2017, Neuroscience
Citation Excerpt :
However, the same analyses revealed tendencies for lower flow during I3 corresponding to filling of the circle of Willis formed by the continuation of the internal carotids and the posterior communicating artery (t1,14 = 1.98, p = 0.067) as well as during I4 all through which the anterior and middle cerebral arteries were filled (t1,14 = 1.92, p = 0.074) in mice that were given LPS as compared to those that received its vehicle. Since blood flow may increase with time of isoflurane anesthesia (Sicard et al., 2003; Duong, 2007; Shin et al., 2007), the relative number of voxels that appeared between sequential images of the filling of the circle of Willis were also analyzed by repeated-measures ANOVAs. These analyses indicated indeed significantly increased flow over time after iv injection (F5,65 = 2.52: p < 0.05), but no effect of treatment during I1 (Fig. 1A), that is during filling of the internal carotids upstream of the circle of Willis and that of the basilar artery.
Brain dysfunction is a frequent complication of the systemic inflammatory response to bacterial infection or sepsis. In the present work, the effects of intravenous bacterial lipopolysaccharide (LPS) administration on cerebral arterial blood flow were assessed with time-of-flight (TOF)-based magnetic resonance angiography (MRA) in mice. Cerebral expression of the transcription factors nuclear factor-kappaB (NF-κB) and c-Fos and that of enzymes synthesizing vasoactive mediators, such as prostaglandins and nitric oxide, known to be increased under inflammatory conditions, were studied in the same animals. Time-resolved TOF MRA revealed no differences in blood flow in the internal carotids upstream of the circle of Willis, but indicated lower flow in its lateral parts as well as in the middle and anterior cerebral arteries after intravenous LPS injection as compared to saline administration. Although LPS did not increase c-Fos expression in ventral forebrain structures of these animals, it did induce NF-κB in meningeal blood vessels. LPS also increased cerebral expression of cyclooxygenase-2 and prostaglandin E synthase mRNAs, but de novo expression occurred in veins rather than in arteries. In conclusion, our work indicates that LPS-induced systemic inflammation does not necessarily affect filling of the circle of the Willis from the periphery, but that circulating LPS alters outflow from the circle of Willis to the middle and anterior cerebral arteries. These modifications in arterial flow were not related to increased cerebral synthesis of prostaglandins, but may instead be the consequence of the action of circulating prostaglandins and other vasoactive mediators on brain-irrigating arteries during systemic inflammation.
Spatiotemporal changes in blood-brain barrier permeability, cerebral blood flow, T<inf>2</inf> and diffusion following mild traumatic brain injury
2016, Brain Research
The blood-brain barrier (BBB) can be impaired following traumatic brain injury (TBI), however the spatiotemporal dynamics of BBB leakage remain incompletely understood. In this study, we evaluated the spatiotemporal evolution of BBB permeability using dynamic contrast-enhanced MRI and measured the volume transfer coefficient (K^trans), a quantitative measure of contrast agent leakage across the blood and extravascular compartment. Measurements were made in a controlled cortical impact (CCI) model of mild TBI in rats from 1 h to 7 days following TBI. The results were compared with cerebral blood flow, T₂ and diffusion MRI from the same animal. Spatially, K^trans changes were localized to superficial cortical layers within a 1 mm thickness, which was dramatically different from the changes in cerebral blood flow, T₂ and diffusion, which were localized to not only the superficial layers but also to brain regions up to 2.2 mm from the cortical surface. Temporally, K^trans changes peaked at day 3, similar to CBF and ADC changes, but differed from T₂ and FA, whose changes peaked on day 2. The pattern of superficial cortical layer localization of K^trans was consistent with patterns revealed by Evans Blue extravasation. Collectively, these results suggest that BBB disruption, edema formation, blood flow disturbance and diffusion changes are related to different components of the mechanical impact, and may play different roles in determining injury progression and tissue fate processes following TBI.

View all citing articles on Scopus

View full text

Research ReportCerebral blood flow and BOLD fMRI responses to hypoxia in awake and anesthetized rats

Abstract

Introduction

Section snippets

Physiological parameters

Discussion

Animal preparation

Acknowledgments

J. Neurosci. Methods

Brain Res.

NeuroImage

Neuron.

Acute blood pressure elevation during repetitive hypocapnic and eucapnic hypoxia in rats

J. Appl. Physiol.

Hypoxia increases velocity of blood flow through parenchymal microvascular systems in rat brain

J. Cereb. Blood Flow Metab.

MR contrast due to intravascular magnetic susceptibility perturbations

Magn. Reson. Med.

Calibrated functional MRI: mapping the dynamics of oxidative metabolism

Proc. Natl. Acad. Sci. U. S. A.

Enhancing fMRI contrast in awake-behaving primates using intravascular magnetite dextran nanoparticles

NeuroReport

Functional MRI of calcium-dependent synaptic activity: cross correlation with CBF and BOLD measurements

Magn. Reson. Med.

Effect of hyperoxia, hypercapnia and hypoxia on cerebral interstitial oxygen tension and cerebral blood flow in the rat brain: an 19F/1H study

Magn. Reson. Med.

Localized blood flow response at sub-millimeter columnar resolution

Proc. Natl. Acad. Sci. U. S. A.

Evidence of the limitations of water as a freely diffusible tracer in brain of the Rhesus monkey

Circ. Res.

The effects of changes in PaCO2 on cerebral blood volume, blood flow, and vascular mean transit time

Stroke

What is the correct value for the brain–blood partition coefficient for water?

J. Cereb. Blood Flow Metab.

Cerebral autoregulation in awake versus isoflurane-anesthetized rats

Anesth. Analg.

Linear coupling between cerebral blood flow and oxygen consumption in activated human cortex

Proc. Natl. Acad. Sci.

Research Report
Cerebral blood flow and BOLD fMRI responses to hypoxia in awake and anesthetized rats