Tadeusz Foniok

MR molecular imaging of early endothelial activation in focal ischemia.

Focal ischemia followed by reperfusion initiates a harmful P‐ and E‐selectin–mediated recruitment of leukocytes in brain microvasculature. In this study, we tested whether a novel magnetic resonance (MR) contrast agent (Gd‐DTPA‐sLex A), which is designed to bind to activated endothelium could be detected by MR imaging (MRI) in a focal stroke mouse model. MRIs (9.4T) of the brain were acquired 24 hours after transient middle cerebral artery occlusion. T1 maps were acquired repeatedly before and up to 1.5 hours after the intravenous injection of either Gd‐DTPA or Gd‐DTPA‐sLex A. Analysis of images included a pixel‐by‐pixel subtraction of T1 maps from the precontrast T1 maps and quantification of T1 within the ischemic area. After injection of Gd‐DTPA‐sLex A, T1 decreased compared with precontrast levels, and an interhemispheric difference between the pre–post contrast T1 developed within the stroke lesion at a mean time of 52 minutes after injection (p < 0.05). Animals injected with Gd‐DTPA did not exhibit changes in T1 signal intensity between regions of the ipsilateral and contralateral hemispheres, indicating that the reductions in T1 observed with Gd‐DTPA‐sLex A were unrelated to blood–brain barrier breakdown. Fluorescent‐labeled sLex A administered intravenously was observed to bind to the endothelium of injured but not control brain. The study suggests that the contrast agent Gd‐DTPA‐sLex A can be used to visualize early endothelial activation after transient focal ischemia in vivo with MRI. Ann Neurol 2004;56:116–120

Functional MRI of the rat lumbar spinal cord involving painful stimulation and the effect of peripheral joint mobilization

To examine neuronal activation in the spinal cord due to secondary hyperalgesia resulting from intrajoint capsaicin injection, and the effect of physiotherapy manipulation, using functional magnetic resonance imaging (fMRI), in α‐chloralose anesthetized rats.

Functional MRI involving painful stimulation of the ankle and the effect of physiotherapy joint mobilization

We examined whether cerebral activation due to secondary hyperalgesia resulting from intrajoint capsaicin injection could be detected using functional magnetic resonance imaging (fMRI) in alpha-chloralose anesthetized rats. We also examined whether we could detect analgesic changes in the central nervous system response to pain as a result of physiotherapy joint manipulation. Robust activation of areas of the brain known to be associated with the processing of pain, namely the anterior cingulate (bilateral), frontal cortex (bilateral) and sensory motor cortex (contralateral), was found in all animals following injection of 25 microl of capsaicin (128 microg/ml in 7.5% DMSO) into the plantar surface of the rat hindpaw (n = 7) and 75 microL into the ankle joint (n = 13). Significantly greater activation was observed when capsaicin was injected into the plantar surface of the hindpaw compared to the ankle joint. Mechanical allodynia and secondary hyperalgesia following capsaicin injection into the ankle joint also resulted in activation of the same brain regions. Trends toward decreased areas of activation in brain regions associated with pain in animals following physiotherapy joint mobilization were observed.

Magnetic resonance imaging of differential gray versus white matter injury following a mild or moderate hypoxic-ischemic insult in neonatal rats

Selective white matter injury in the pre-mature infants suggests it has a greater susceptibility to hypoxia-ischemia. To investigate whether white matter injury would predominate following a mild hypoxic-ischemic insult, 7-day-old rats underwent either mild or moderate hypoxia-ischemia and magnetic resonance imaging 24 h later. Mild and moderate hypoxia-ischemia were produced by unilateral carotid artery occlusion plus exposure to hypoxia for either 45-50 or 90 min at ambient temperatures of 34.5 or 35.5 degrees C, respectively. Following mild hypoxia-ischemia, there was a significant increase in T(1) and T(2) within periventricular white matter (e.g. corpus callosum) in the hemisphere ipsilateral to the occlusion compared to that contralaterally and less of an increase within gray matter (e.g. cortex and striatum). This corresponded to relatively selective white matter injury detected histologically. Following a moderate hypoxia-ischemia, both gray and white matter was severely injured with marked increases in T(1) and T(2) occurring in both white and gray matter regions ipsilateral to the hypoxia-ischemia. We conclude that a mild insult, consisting of a short duration of hypoxia-ischemia at a slightly lower body temperature than a moderate hypoxic-ischemic insult, produces enhanced injury in white matter and a relative sparing of gray matter.

Simultaneous functional magnetic resonance imaging in the rat spinal cord and brain.

Functional magnetic resonance imaging (fMRI) method was developed to investigate the pattern and temporal relationship in neuronal pathways of brain and spinal cord. Signal intensity changes correlating with stimulation patterns were observed simultaneously in the rat spinal cord and brain using fMRI at 9.4 T. Electrical stimulation of the forepaw was used to elicit activity. A quadrature volume RF coil covering both brain and the cervical spinal cord was used. Sets of fast spin echo (FSE) images were acquire simultaneously for both brain and spinal cord fMRI. Experiments were repeated in single animal and across animals. Activities within the dorsal horn of the spinal cord and within the somatosensory cortex were observed consistently within each animal as well as across animals.

Cellular correlates of longitudinal diffusion tensor imaging of axonal degeneration following hypoxic-ischemic cerebral infarction in neonatal rats

Ischemically damaged brain can be accompanied by secondary degeneration of associated axonal connections e.g. Wallerian degeneration. Diffusion tensor imaging (DTI) is widely used to investigate axonal injury but the cellular correlates of many of the degenerative changes remain speculative. We investigated the relationship of DTI of directly damaged cerebral cortex and secondary axonal degeneration in the cerebral peduncle with cellular alterations in pan-axonal neurofilament staining, myelination, reactive astrocytes, activation of microglia/macrophages and neuronal cell death. DTI measures (axial, radial and mean diffusivity, and fractional anisotropy (FA)) were acquired at hyperacute (3 h), acute (1 and 2 d) and chronic (1 and 4 week) times after transient cerebral hypoxia with unilateral ischemia in neonatal rats. The tissue pathology underlying ischemic and degenerative responses had a complex relationship with DTI parameters. DTI changes at hyperacute and subacute times were smaller in magnitude and tended to be transient and/or delayed in cerebral peduncle compared to cerebral cortex. In cerebral peduncle by 1 d post-insult, there were reductions in neurofilament staining corresponding with decreases in parallel diffusivity which were more sensitive than mean diffusivity in detecting axonal changes. Ipsilesional reductions in FA within cerebral peduncle were robust in detecting both early and chronic degenerative responses. At one or four weeks post-insult, radial diffusivity was increased ipsilaterally in the cerebral peduncle corresponding to pathological evidence of a lack of ontogenic myelination in this region. The detailed differences in progression and magnitude of DTI and histological changes reported provide a reference for identifying the potential contribution of various cellular responses to FA, and, parallel, radial, and mean diffusivity.

Imaging Corticospinal Degeneration in Neonatal Rats with Unilateral Cerebral Infarction

Recent human studies indicate that magnetic resonance (MR) imaging, particularly diffusion weighted imaging, detects abnormalities within the descending cortico-spinal tract following stroke. Whether these changes are directly related to processes of axonal degeneration and how MR changes (e.g. apparent diffusion coefficient of water (ADC) and T(2)) vary in their diagnostic utility over time is not known. The present study demonstrates that a commonly used rat model of neonatal transient unilateral hypoxia-ischemia provides similar diffusion weighted and ADC changes in the cerebral peduncle as those observed in human neonates clinically. Imaging the descending cortico-spinal tract in this model at defined acute (1-3 days) and chronic (1 and 4 weeks) time points demonstrates increased T(2) and progressive changes in ADC within the descending cortico-spinal tract in the first days to weeks following hypoxia-ischemia with a normalization by 1 week and further increases in ispilateral cerebral cortex by 4 weeks. These imaging changes are associated with reduced axonal neurofilament staining both at the subacute and more chronic time points. This demonstrates directly the utility of ADC and T(2) MRI to detect acute changes in axons associated with early Wallerian degeneration.

Mild cerebral hypoxia-ischemia produces a sub-acute transient inflammatory response that is less selective and prolonged after a substantial insult

Cerebral ischemia initiates various injurious processes including neuroinflammatory responses such as activation of microglia and increases in cytokine and nitric oxide release. Evidence primarily from in vitro studies, indicates that neuroinflammatory effects can be either beneficial or harmful, possibly related to stimulus strength. We investigated using in vivo models, the effect of a mild or substantial cerebral hypoxia–ischemia on: cerebral microglial/macrophage activation (ED1), pro‐inflammatory cytokines (tumor necrosis factor‐alpha), nitrosative stress (nitrotyrosine) and permanent brain damage. A mild insult produced a transient (1–2 days post) increase in activated microglia/macrophages within subcortical white and not gray matter but transiently increased cytokine or nitrotyrosine expression in cortex and not white matter. There was also prolonged scattered cell death in cortex and white matter over weeks along with loss of myelin/axons and cortical atrophy at 4 weeks post‐insult. In contrast, a substantial insult produced white and gray matter necrosis, cyst formation and atrophy, along with increases in tumor necrosis factor and nitrotyrosine staining within both white and gray matter starting at 1–2 days post‐insult. Microglial/macrophage staining was increased starting at 1‐week post a substantial insult and remained elevated for weeks thereafter.

Cerebral blood flow response to a hypoxic-ischemic insult differs in neonatal and juvenile rats.

To compare the cerebral blood flow (CBF) response to a transient episode of hypoxia–ischemia producing damage in neonatal and juvenile rats. One- and four-week-old rats were subjected to unilateral carotid artery occlusion plus hypoxia (8% oxygen). Perfusion MR images were acquired either in sham controls or in hypoxic–ischemic rats before, during, 1 h and 24 h after hypoxia–ischemia. At 24 h post hypoxia–ischemia, T2 maps and histology were used to assess damage. In sham controls, CBF increased twofold between the age of one and four weeks. Reductions in CBF ipsilateral to the occlusion occurred during hypoxia–ischemia followed by a substantial recovery at 1 h post in both age groups. However, contralaterally, hyperemia occurred during hypoxia–ischemia in four-week but not one-week-old rats. Similarly, hyperemia occurred ipsilaterally at 24 h post hypoxia–ischemia in four-week but not one-week-olds, corresponding to the distribution of elevations in T2. Despite CBF differences, extensive cell death occurred ipsilaterally in both age groups. The CBF responses to hypoxia–ischemia and reperfusion differ depending on postnatal age, with hyperemia occurring in juvenile but not neonatal rats. The results suggest a greater CBF responsiveness and differential relationship between post-ischemic vascular perfusion and tissue injury in older compared with immature animals.

Functional magnetic resonance imaging within the rat spinal cord following peripheral nerve injury.

Functional magnetic resonance imaging (fMRI) was used to detect the effects of graded peripheral nerve injury at the spinal level. Graded peripheral nerve injury in rats was accomplished by transection of nerves entering the spinal cord at the L3 and L4 levels of the spinal cord segments. Electrical stimulation of the hindpaw was used to elicit activity within the spinal cord. The stimulation experimental paradigm consisted of 62 functional images, 5 slices each, with a total of 3 rest and 2 stimulation periods. A 9.4 T MRI system and a quadrature volume rf coil covering the lumbar spinal cord were used for the fMRI study. Sets of fast spin echo images were acquired repeatedly following sham preparatory surgery under control conditions and in rats following sham surgery (pre nerve cut), followed by L3 nerve and then L4 nerve section. In rats with sham surgery, there was a significant activation within the dorsal horn of slices corresponding to L3 and L4 spinal cord segments. Following section of the L3 nerve, there was a reduction in the number of active voxels in the L3 and L4 spinal cord segments. The activation was reduced further by sectioning of the L4 nerve. Thus, following an increasing loss of axonal connections to the spinal cord, there was a decreasing number of active voxels within the spinal cord. The results demonstrate that spinal fMRI in the rat has sufficient sensitivity to detect within the spinal cord the effects of a graded reduction in peripheral connectivity.