Markus Rudin

Molecular imaging in drug discovery and development

Imaging sciences have grown exponentially during the past three decades, and many techniques, such as magnetic resonance imaging, nuclear tomographic imaging and X-ray computed tomography, have become indispensable in clinical use. Advances in imaging technologies and imaging probes for humans and for small animals are now extending the applications of imaging further into drug discovery and development, and have the potential to considerably accelerate the process. This review summarizes some of the recent developments in conventional and molecular imaging, and highlights their impact on drug discovery.

Monocyte chemoattractant protein-1 deficiency is protective in a murine stroke model

Inflammatory processes have been implicated in the pathogenesis of brain damage after stroke. In rodent stroke models, focal ischemia induces several proinflammatory chemokines, including monocyte chemoattractant protein-1 (MCP-1). The individual contribution to ischemic tissue damage, however, is largely unknown. To address this question, the authors subjected MCP-1-deficient mice (MCP-1−/−) to permanent middle cerebral artery occlusion (MCAO). Measurement of basal blood pressure, cerebral blood flow, and blood volume revealed no differences between wild-type (wt) and MCP-1−/− mice. MCAO led to similar cerebral perfusion deficits in wt and MCP-1−/− mice, excluding differences in the MCA supply territory and collaterals. However, compared with wt mice, the mean infarct volume was 29% smaller in MCP-1−/− mice 24 hours after MCAO (P = 0.022). Immunostaining showed a reduction of phagocytic macrophage accumulation within infarcts and the infarct border in MCP-1−/− mice 2 weeks after MCAO. At the same time point, the authors found an attenuation of astrocytic hypertrophy in the infarct border and thalamus in MCP-1−/− mice. However, these effects on macrophages and astrocytes in MCP-1−/− mice occurred too late to suggest a protective role in acute infarct growth. Of note: at 6 hours after MCAO, MCP-1−/− mice produced significantly less interleukin-1β in ischemic tissue; this might be related to tissue protection. The results of this study indicate that inhibition of MCP-1 signaling could be a new acute treatment approach to limit infarct size after stroke.

In vivo detection of amyloid-β deposits by near-infrared imaging using an oxazine-derivative probe

As Alzheimers disease pathogenesis is associated with the formation of insoluble aggregates of amyloid β-peptide, approaches allowing the direct, noninvasive visualization of plaque growth in vivo would be beneficial for biomedical research. Here we describe the synthesis and characterization of the near-infrared fluorescence oxazine dye AOI987, which readily penetrates the intact blood-brain barrier and binds to amyloid plaques. Using near-infrared fluorescence imaging, we demonstrated specific interaction of AOI987 with amyloid plaques in APP23 transgenic mice in vivo, as confirmed by postmortem analysis of brain slices. Quantitative analysis revealed increasing fluorescence signal intensity with increasing plaque load of the animals, and significant binding of AOI987 was observed for APP23 transgenic mice aged 9 months and older. Thus, AOI987 is an attractive probe to noninvasively monitor disease progression in animal models of Alzheimer disease and to evaluate effects of potential Alzheimer disease drugs on the plaque load.

MRI‐based monitoring of inflammation and tissue damage in acute and chronic relapsing EAE

Experimental autoimmune encephalomyelitis (EAE) is a commonly used animal model that in several respects mimics human multiple sclerosis (MS), and can be used to design or validate new strategies for treatment of this disease. In the present study, different MRI techniques (macrophage tracking based on labeling cells in vivo by ultrasmall particles of iron oxide (USPIO), blood–brain barrier (BBB) breakdown, and magnetization transfer imaging (MTI)), as well as immunohistological staining were used to study the burden of disease in Lewis rats immunized by guinea pig myelin. The resulting imaging data was compared with behavioral readouts. Animals were studied during the acute phase and the first relapse. Activated monocytes were detected during both episodes in the brain stem or cortex. These areas coincided in part with areas of BBB breakdown. Significant changes of the magnetization transfer ratios (MTRs) of up to 35% were observed in areas of USPIO accumulation. This suggests that infiltrating monocytes are the major source of demyelination in EAE, but monocyte infiltration and breakdown of the BBB are temporally or spatially independent inflammatory processes. Magn Reson Med 50:309–314, 2003.

Dynamic patterns of USPIO enhancement can be observed in macrophages after ischemic brain damage

Cells of the mononuclear phagocytotic system (MPS) are often found near to or within ischemic tissue and can potentially aggravate cellular damage. Hence, visualization of those cells would allow demarcation of putatively affected from intact tissue. Experimental MRI studies have shown that ultrasmall particles of dextran‐coated iron oxide (USPIO) are internalized into cells of the MPS. To test if this cell tagging method may be also applied to cerebral infarction, USPIOs were administered to Fisher rats 5.5 h after permanent occlusion of the middle cerebral artery (pMCAO). During the first 2 days USPIO were preferentially found in patches within the lesion and in surrounding areas. On day 4, USPIOs expanded within the core of the lesion. On day 7 they were found predominantly within the boundary area. Histological analysis showed large populations of macrophages containing iron particles in the infarcted tissue. We conclude, therefore, that it is possible to monitor MPS activity after focal cerebral ischemia using USPIOs. Magn Reson Med 46:1018–1022, 2001.

Functional and Anatomical Reorganization of the Sensory-Motor Cortex after Incomplete Spinal Cord Injury in Adult Rats

A lateral hemisection injury of the cervical spinal cord results in Brown-Séquard syndrome in humans and rats. The hands/forelimbs on the injured side are rendered permanently impaired, but the legs/hindlimbs recover locomotor functions. This is accompanied by increased use of the forelimb on the uninjured side. Nothing is known about the cortical circuits that correspond to these behavioral adaptations. In this study, on adult rats with cervical spinal cord lateral hemisection lesions (at segment C3/4), we explored the sensory representation and corticospinal projection of the intact (ipsilesional) cortex. Using blood oxygenation level-dependent functional magnetic resonance imaging and voltage-sensitive dye (VSD) imaging, we found that the cortex develops an enhanced representation of the unimpaired forepaw by 12 weeks after injury. VSD imaging also revealed the cortical spatio-temporal dynamics in response to electrical stimulation of the ipsilateral forepaw or hindpaw. Interestingly, stimulation of the ipsilesional hindpaw at 12 weeks showed a distinct activation of the hindlimb area in the intact, ipsilateral cortex, probably via the injury-spared spinothalamic pathway. Anterograde tracing of corticospinal axons from the intact cortex showed sprouting to recross the midline, innervating the spinal segments below the injury in both cervical and lumbar segments. Retrograde tracing of these midline-crossing axons from the cervical spinal cord (at segment C6/7) revealed the formation of a new ipsilateral forelimb representation in the cortex. Our results demonstrate profound reorganizations of the intact sensory-motor cortex after unilateral spinal cord injury. These changes may contribute to the behavioral adaptations, notably for the recovery of the ipsilesional hindlimb.

Predictability of FTY720 efficacy in experimental autoimmune encephalomyelitis by in vivo macrophage tracking: Clinical implications for ultrasmall superparamagnetic iron oxide-enhanced magnetic resonance imaging

To examine the efficacy of FTY720 as a new agent to reduce inflammatory activity in an animal model of multiple sclerosis (MS) by in vivo macrophage tracking.

Rewiring of hindlimb corticospinal neurons after spinal cord injury

Little is known about the functional role of axotomized cortical neurons that survive spinal cord injury. Large thoracic spinal cord injuries in adult rats result in impairments of hindlimb function. Using retrograde tracers, we found that axotomized corticospinal axons from the hindlimb sensorimotor cortex sprouted in the cervical spinal cord. Mapping of these neurons revealed the emergence of a new forelimb corticospinal projection from the rostral part of the former hindlimb cortex. Voltage-sensitive dye (VSD) imaging and blood-oxygen-level–dependent functional magnetic resonance imaging (BOLD fMRI) revealed a stable expansion of the forelimb sensory map, covering in particular the former hindlimb cortex containing the rewired neurons. Therefore, axotomised hindlimb corticospinal neurons can be incorporated into the sensorimotor circuits of the unaffected forelimb.

Recovery and brain reorganization after stroke in adult and aged rats

Stroke is a prevalent and devastating disorder, and no treatment is currently available to restore lost neuronal function after stroke. One unique therapy that improves recovery after stroke is neutralization of the neurite inhibitory protein Nogo‐A. Here, we show, in a clinically relevant model, improved functional recovery and brain reorganization in the aged and adult rat when delayed anti–Nogo‐A therapy is given after ischemic injury. These results support the efficacy of Nogo‐A neutralization as treatment for ischemic stroke, even in the aged animal and after a 1‐week delay, and implicate neuronal plasticity from unlesioned areas of the central nervous system as a mechanism for recovery. Ann Neurol 2005;58:950–953

Simultaneous BOLD fMRI and fiber-optic calcium recording in rat neocortex

Functional magnetic resonance imaging (fMRI) based on blood oxygen level–dependent (BOLD) contrast is widely used for probing brain activity, but its relationship to underlying neural activity remains elusive. Here, we combined fMRI with fiber-optic recordings of fluorescent calcium indicator signals to investigate this relationship in rat somatosensory cortex. Electrical forepaw stimulation (1–10 Hz) evoked fast calcium signals of neuronal origin that showed frequency-dependent adaptation. Additionally, slower calcium signals occurred in astrocyte networks, as verified by astrocyte-specific staining and two-photon microscopy. Without apparent glia activation, we could predict BOLD responses well from simultaneously recorded fiber-optic signals, assuming an impulse response function and taking into account neuronal adaptation. In cases with glia activation, we uncovered additional prolonged BOLD signal components. Our findings highlight the complexity of fMRI BOLD signals, involving both neuronal and glial activity. Combined fMRI and fiber-optic recordings should help to clarify cellular mechanisms underlying BOLD signals.