Neurological and behavioral abnormalities, ventricular dilatation, altered cellular functions, inflammation, and neuronal injury in brains of mice due to common, persistent, parasitic infection.

BACKGROUND: Worldwide, approximately two billion people are chronically infected with Toxoplasma gondii with largely unknown consequences. METHODS: To better understand long-term effects and pathogenesis of this common, persistent brain infection, mice were infected at a time in human years equivalent to early to mid adulthood and studied 5-12 months later. Appearance, behavior, neurologic function and brain MRIs were studied. Additional analyses of pathogenesis included: correlation of brain weight and neurologic findings; histopathology focusing on brain regions; full genome microarrays; immunohistochemistry characterizing inflammatory cells; determination of presence of tachyzoites and bradyzoites; electron microscopy; and study of markers of inflammation in serum. Histopathology in genetically resistant mice and cytokine and NRAMP knockout mice, effects of inoculation of isolated parasites, and treatment with sulfadiazine or alphaPD1 ligand were studied. RESULTS: Twelve months after infection, a time equivalent to middle to early elderly ages, mice had behavioral and neurological deficits, and brain MRIs showed mild to moderate ventricular dilatation. Lower brain weight correlated with greater magnitude of neurologic abnormalities and inflammation. Full genome microarrays of brains reflected inflammation causing neuronal damage (Gfap), effects on host cell protein processing (ubiquitin ligase), synapse remodeling (Complement 1q), and also increased expression of PD-1L (a ligand that allows persistent LCMV brain infection) and CD 36 (a fatty acid translocase and oxidized LDL receptor that mediates innate immune response to beta amyloid which is associated with pro-inflammation in Alzheimer's disease). Immunostaining detected no inflammation around intra-neuronal cysts, practically no free tachyzoites, and only rare bradyzoites. Nonetheless, there were perivascular, leptomeningeal inflammatory cells, particularly contiguous to the aqueduct of Sylvius and hippocampus, CD4+ and CD8+ T cells, and activated microglia in perivascular areas and brain parenchyma. Genetically resistant, chronically infected mice had substantially less inflammation. CONCLUSION: In outbred mice, chronic, adult acquired T. gondii infection causes neurologic and behavioral abnormalities secondary to inflammation and loss of brain parenchyma. Perivascular inflammation is prominent particularly contiguous to the aqueduct of Sylvius and hippocampus. Even resistant mice have perivascular inflammation. This mouse model of chronic T. gondii infection raises questions of whether persistence of this parasite in brain can cause inflammation or neurodegeneration in genetically susceptible hosts.

Use of Microcomputed Tomography to Measure the Relaxin-induced Expansion of Intrapubic Ligaments in Mice.

Relaxin is a 6-kDa peptide in the insulin superfamily of hormones. In addition to its effects on reproductive and musculoskeletal ligaments, relaxin has demonstrated beneficial effects on cardiac, renal, and vascular systems in preclinical models. The mouse intrapubic ligament ex vivo bioassay is the current standard for measuring in vivo relaxin bioactivity. However, this bioassay necessitates euthanasia and dissection of large cohorts to measure the intrapubic ligament at specified time points. We hypothesized that µCT imaging could be used to reduce the number of animals necessary for the intrapubic ligament bioassay by enabling a single animal to be followed longitudinally throughout the study rather than euthanizing different cohorts at established time points. Female CD1 mice were used to compare µCT imaging with the current standard. Both protocols revealed significant differences in intrapubic ligament length, with the µCT data having greater power when corrected for baseline imaging. From these data, we concluded that using µCT to measure the intrapubic ligament in mice primed with estrogen and dosed with relaxin is a viable refinement and will allow the use of fewer animals in longitudinal studies and provide more robust data, because animals can serve as their own controls.

Increases in oxygen consumption without cerebral blood volume change during visual stimulation under hypotension condition.

The magnitude of the blood oxygenation level-dependent (BOLD) signal depends on cerebral blood flow (CBF), cerebral blood volume (CBV) and cerebral metabolic rate of oxygen (CMRO2). Thus, it is difficult to separate CMRO2 changes from CBF and CBV changes. To detect the BOLD signal changes induced only by CMRO2 responses without significant evoked CBF and CBV changes, BOLD and CBV functional magnetic resonance imaging (fMRI) responses to visual stimulation were measured under normal and hypotension conditions in isoflurane-anesthetized cats at 4.7 T. When the mean arterial blood pressure (MABP) decreased from 89+/-10 to 50+/-1 mm Hg (mean+/-standard deviation, n=5) by infusion of vasodilator sodium nitroprusside, baseline CBV in the visual cortex increased by 28.4%+/-8.3%. The neural activity-evoked CBV increase in the visual cortex was 10.8%+/-3.9% at normal MABP, but was negligible at hypotension. Positive BOLD changes of +1.8%+/-0.5% (gradient echo time=25 ms) at normal MABP condition became prolonged negative changes of -1.2%+/-0.3% at hypotension. The negative BOLD response at hypotension starts approximately 1 sec earlier than positive BOLD response, but similar to CBV change at normal MABP condition. Our finding shows that the negative BOLD signals in an absence of CBV changes are indicative of an increase in CMRO2. The vasodilator-induced hypotension model simplifies the physiological source of the BOLD fMRI signals, providing an insight into spatial and temporal CMRO2 changes.

Endothelin in a murine model of cerebral malaria.

Cerebral malaria (CM) remains a deadly complication of Plasmodium falciparum infection, and children are at high risk of developing encephalopathy as a result of CM. This is probably a consequence of the activation of many of the inflammatory cytokines as well as the glial cells and the vascular endothelium in the brain. We have previously demonstrated that there is a striking reduction in cerebral blood flow by magnetic resonance imaging when mice are infected with Plasmodium berghei ANKA (PbA), and we now demonstrate a possible role for endothelin (ET-1) in the pathogenesis of CM. The brains of female C57BL/6 mice with PbA infection were examined at Day 5 for the expression of ET-1, endothelin converting enzyme (ECE), and the endothelin receptors A and B (ET(A) and ET(B)) by both reverse transcription-polymerase chain reaction (RT-PCR) and quantitative real-time PCR. ET-1 and ECE mRNA expression was markedly increased by RT-PCR in PbA-infected mice. Real-time quantitative PCR demonstrated a 3-fold increase in ET-1 (P < 0.05) and a significant increase in ET(A) and ET(B) expression (P < 0.05) in PbA-infected mice. Histopathology bof PbA-infected mice demonstrated a transformation in the morphology of microglial cells and clustering of these cells consistent with activation. Though the full impact of ET-1 on CM remains to be elucidated, these findings demonstrate that in the murine model, there is a significant increase in ET-1 and its components, which is associated with the vasculopathy and immunopathology of CM.

Sensitivity enhancement and compensation of RF penetration artifact with planar actively detunable quadrature surface coil.

An actively detunable planar quadrature surface coil for human body imaging at 4 T has been constructed and compared with a conventional linear surface coil. The coil could be used as a transmit/receive or a receive-only device in combination with a volume transmit coil. Transmission, reception profiles and the corresponding images acquired with each coil, as well as with both individual modes of the quadrature coil, are presented. Data collected using a tissue equivalent loaded phantom recorded with the linear surface coil demonstrated significant intensity distortions due to RF penetration artifact. The quadrature surface coil, on the other hand, provided compensation of the artifact, separately in its transmission and reception profiles as well as in the resultant images. Substantial sensitivity gain was also observed for the quadrature coil compared to the linear device. Significant advantages of using the quadrature surface coil over the linear device at 4 T have, therefore, been demonstrated.

Human cerebral blood flow and metabolism in acute insulin-induced hypoglycemia.

How the human brain functions under conditions of acute hypoglycemia remains a complex question by virtue of the potential simultaneous shifts in processes of perfusion, metabolism, and changing demand. We examined this issue by measuring cerebral blood flow (CBF) and oxidative metabolism (CMRO2) in insulin-induced hypoglycemic (HG) and euglycemic (EG) conditions at rest and during motor activation in normal human subjects using magnetic resonance (MR). Experiments were performed on 12 subjects (9M, 3F). The protocol consisted of insulin-induced hypoglycemia (targeting a HG of 60 mg/dL) followed by euglycemia, or in reverse order, each phase lasting approximately 1.5 h. Euglycemia was performed with the same insulin infusion rate so as to match the hypoglycemic phase. Magnetic resonance data were acquired 30 mins after the target plasma glucose was achieved so as to minimize any acute effects. Although the depth of hypoglycemia achieved in the present study was relatively small, the present data found a significant increase in flow in motor cortex with mild hypoglycemia, from 56.4+/-13.6 mL/100 g min (euglycemia) to 64.3+/-7.6 mL/100 g min (hypoglycemia). Using the Renkin-Crone exponential model of oxygen extraction with MR models of susceptibility-based relaxation, analysis of the flow measurements, relaxation and BOLD data also implied that throughout the studies, metabolism and flow remained coupled. Elementary motor task activation was not associated with any consistent larger activated flows. Thus it remains that although mild hypoglycemia induced an increase in basal flow and metabolism, a similar increase was not seen in task activation.

Continuous-wave near-infrared spectroscopy using pathlength-independent hypoxia normalization.

A general physiological model for the hemodynamic response during altered blood flow, oxygenation, and metabolism is presented. Calculations of oxy-, deoxy-, and total hemoglobin changes during stimulation are given. It is shown that by using a global hyperoxic or mild hypoxic challenge it is possible to normalize the activation response in terms of the fractional changes in the cerebral blood volume, tissue oxygenation index, and oxygen extraction ratio, which are independent of the optical pathlength. Using a dual wavelength spectrometer, the method is validated by measuring pathlength-independent hemodynamic responses during mild hypercarbia in a rat model. Phantom experiments showed that the changes in optical pathlength were small as the hemoglobin concentration was varied over a wide range. The determination of quantitative parameters facilitates the use of continuous-wave transcranial methods by providing a means by which to characterize activation response across subjects.

Dominant events that modulate mass transfer coefficient of oxygen in cerebral cortex.

Recently, a model of cerebral oxygen delivery was described (J Appl Physiol 85:554) which yields a relationship similar to that used to depict substrate transport across the endothelium. Because the endothelium is not a diffusion barrier for oxygen, the permeability surface area product was replaced by an effective mass transfer coefficient term for oxygen, D. The cerebral metabolic rate of oxygen utilization (CMRO2) was linked to cerebral blood flow (CBF) and volume (CBV) through properties that modify the vessel-to-tissue oxygen tension giving rise to changes in D. Changes in the value of D were correlated with changes in CBF, CMRO2, and CBV as measured using NMR methods in a 48 microL volume of the cerebral cortex of anesthetized rats at different levels of activity. We conclude that the changes in total vascular volume (i.e., swelling or shrinking of the capillary bed) contributes < 5% to changes in D, whereas variations in the number of hematic vs. plasmatic capillaries, or intra-capillary stacking vs. unpacking of erythrocytes, or increase vs. decrease of dissolved oxygen in the tissue (i.e., processes which modify vessel-to-tissue oxygen tension) contribute(s) > 95% to changes in D.

SENSE imaging with a quadrature half-volume transverse electromagnetic (TEM) coil at 4T.

PURPOSE: To demonstrate the feasibility of high-field SENSE imaging of large objects, such as the human head, using a semicircular (half-volume) coil for both transmission and multi-channel reception. MATERIALS AND METHODS: As a proof of concept, we present experimental data obtained using a seven-element half-volume (180 degrees of arc) transmit/receive quadrature transverse electromagnetic (TEM) coil. SENSE images of the human brain were acquired with a reduction factor of R=2, using two degenerate linear modes of the same coil as independent receive channels at 4T. Since the need for additional hardware (i.e., a separate set of receive coils) is eliminated, the design can be substantially simplified. RESULTS: The experimental data demonstrate that linear modes of the half-volume TEM coil have essentially no noise correlation, and their sensitivity profiles satisfy the requirement for small g-factors. Also, this type of coil provides efficient transmission with a relatively large uniform region and a reception profile that is more uniform than that of the surface coils. CONCLUSION: We demonstrate the feasibility of SENSE imaging using a half-volume coil. Half-volume coils allow reduced total power deposition compared to full-volume coils, and may replace the latter in body imaging applications in which the target region of interest (ROI) is smaller than the entire torso.

Reduced cerebral blood flow and N-acetyl aspartate in a murine model of cerebral malaria.

Cerebral malaria is an important cause of morbidity and mortality in many parts of the world. It has been suggested that cerebral malaria is associated with reduced perfusion due to the blockage of blood vessels by parasitized erythrocytes; although, no quantitative validation of this has been done. We infected C57BL/6 mice with the ANKA strain of Plasmodium berghei and on day 6 of infection we investigated alterations in brain function using arterial spin labeling MRI and proton MRS. MR images did not demonstrate signs of damage. However, there was a significant reduction in cerebral blood flow (P<0.012) and the ratio of N-acetyl-aspartate (NAA) to creatine (Cr) (P<0.01) relative to non-infected mice. The NAA/Cr ratios were significantly correlated with cerebral perfusion (r=0.87) suggesting a relationship between impaired oxygen delivery and neuronal dysfunction. Pathological examination revealed accumulations of damaged axons providing a correlate for the decreased NAA/Cr ratio in infected mice. This murine model will permit non-invasive studies of neurologic function during malarial infection.

Non-invasive assessment of language lateralization by transcranial near infrared optical topography and functional MRI.

Near infrared optical topography (OT) is the simultaneous acquisition of hemoglobin absorption from an array of optical fibers on the scalp to construct maps of cortical activity. We demonstrate that OT can be used to determine lateralization of prefrontal areas to a language task that has been validated by functional MRI (fMRI). Studies were performed on six subjects using a visually presented language task. Laterality was quantified by the relative number of activated pixels in each hemisphere for fMRI, and the total hemoglobin responses in each hemisphere for OT. All subjects showed varying degrees of left hemisphere language dominance and the mean laterality indices for subjects who underwent both OT and fMRI were in good agreement. These studies demonstrate that OT gives predictions of hemispheric dominance that are consistent with fMRI. Due to the ease of use and portable nature of OT, it is anticipated that optical topography will be valuable tool for neurological examinations of cognitive function.

Decreased cerebral perfusion correlates with increased BOLD hyperoxia response in transgenic mouse models of sickle cell disease.

Neurological complications such as stroke are known consequences of sickle cell disease (SCD). In order to improve methods for the evaluation of stroke risk in SCD, MRI was used to evaluate cerebrovascular function in transgenic mouse models of human SCD. It is hypothesized that oxygen-sensitive imaging in the brain will reveal areas of excess deoxygenation that are either at risk of or the result of vaso-occlusion. Arterial spin labeling (ASL) perfusion was performed in order to correlate BOLD results with microvascular cerebral blood flow. Upon comparison with control animals, there was a relative increase in BOLD hyperoxia response of 42-67% (P < 0.001) in the transgenic mice while cerebral blood flow during normoxia was reduced by 30-40% (P < 0.02). Hyperoxia caused cerebral blood flow to decrease in control mice, whereas blood flow increased in the sickle transgenic mice. These results indicate impairment in brain autoregulation in the sickle cell transgenic mice leading to increased cerebral deoxyhemoglobin. Increased deoxyhemoglobin coupled with reduced perfusion may further increase the risk of vaso-occlusion and stroke. This may reflect polymer reduction or reduced cell adhesion during hyperoxia. The MRI protocol is noninvasive and thus directly applicable to a clinical population.

Simultaneous recording of event-related auditory oddball response using transcranial near infrared optical topography and surface EEG.

Near infrared optical topography (OT) is the measurement of hemoglobin absorption simultaneously from an array of optical fibers on the scalp to construct maps of cortical activity. We demonstrate that OT can be used to simultaneously detect and characterize the hemodynamic responses associated with an "oddball" auditory stimulus and that corresponding electrical event related potentials can be acquired simultaneously using conventional scalp recordings. In addition to the measured electrical response, the hemodynamic localization is consistent with fMRI studies, which show significant activation in the temporal and parietal cortical regions. The event-related response of total hemoglobin showed relatively slow peak latencies (5.8 +/- 0.3 s), which were also consistent with fMRI. The current study shows the regions of peak hemodynamic activity that are in closest proximity to areas of peak electrical activity. This is the first demonstration of simultaneous ERP electrical recording and non-invasive optical mapping in human subjects, which promises to be an important tool in the characterization of both normal and abnormal brain function.