Changes in regional brain perfusion during functional brain activation: comparison of [(64)Cu]-PTSM with [(14)C]-Iodoantipyrine.

A dilemma in behavioral brain mapping is that conventional techniques immobilize the subject, extinguishing all but the simplest behaviors. This is avoided if brain activation is imaged after completion of the behavior and tissue capture of the tracer. A single-pass flow tracer proposed for positron emission tomography (PET) is a radiolabeled copper(II) complex of pyruvaldehyde bis(N(4)-methylthiosemicarbazone), [Cu(64)]-PTSM. [Cu(64)]-PTSM reaches steady-state cerebral distribution more rapidly than the metabolic tracer [(18)F]-fluorodeoxyglucose, allowing imaging with substantially greater temporal resolution. Using dual-label autoradiography, this study compares the relative regional cerebral blood flow tracer distribution (CBF-TR) of [(64)Cu]-PTSM to that of the classic perfusion tracer [(14)C]-iodoantipyrine in a rat model during treadmill walking. Rats were exposed to continuous walking on a treadmill and compared to quiescent controls. [(64)Cu]-PTSM was bolus injected (iv) after 1 min, followed by a 5-minute uptake and subsequent bolus injection of [(14)C]-iodoantipyrine. CBF-TR was quantified by autoradiography and analyzed in the three-dimensionally reconstructed brain by statistical parametric mapping, as well as by region-of-interest analysis. A high homology was found between the [(64)Cu]-PTSM and [(14)C]-iodoantipyrine patterns of cerebral activation in cortical and subcortical regions. For white matter, however, [(64)Cu]-PTSM showed lower perfusion than [(14)Cu]-iodoantipyrine. [(64)Cu]-PTSM is a useful tracer for functional brain mapping in freely-moving subjects. Its application in conjunction with PET promises to increase our understanding of the neural circuitry of behaviors dependent on locomotion.

Brain maps on the go: functional imaging during motor challenge in animals.

Brain mapping in the freely moving animal is useful for studying motor circuits, not only because it avoids the potential confound of sedation or restraints, but because activated brain states may serve to accentuate differences that only manifest partially while a subject is in the resting state. Perfusion or metabolic mapping using autoradiography allows one to examine changes in brain function at the circuit level across the entire brain with a spatial resolution (approximately 100 micro) appropriate for the rat or mouse brain, and a temporal resolution (seconds-minutes) sufficient for capturing acute brain changes. Here we summarize the application of these methods to the functional brain mapping of behaviors involving locomotion of small animals, methods for the three-dimensional reconstruction of the brain from autoradiographic sections, voxel based analysis of the whole brain, and generation of maps of the flattened rat cortex. Application of these methods in animal models promises utility in improving our understanding of motor function in the normal brain, and of the effects of neuropathology and treatment interventions such as exercise have on the reorganization of motor circuits.

Flattened cortical maps of cerebral function in the rat: a region-of-interest approach to data sampling, analysis and display.

We describe a method for the measurement, analysis and display of cerebral cortical data obtained from coronal brain sections of the adult rat. In this method, regions-of-interest (ROI) are selected in the cortical mantle in a semiautomated fashion using a radial grid overlay, spaced in 15 degrees intervals from the midline. ROI measurements of intensity are mapped on a flattened two-dimensional surface. Topographic maps of statistical significance at each ROI allow for the rapid viewing of group differences. Cortical z-scores are displayed with the boundaries of brain regions defined according to a standard atlas of the rat brain. This method and accompanying software implementation (Matlab, Labview) allow for compact data display in a variety of autoradiographic and histologic studies of the structure and function of the rat brain.

Reorganization of functional brain maps after exercise training: Importance of cerebellar-thalamic-cortical pathway.

Exercise training (ET) causes functional and morphologic changes in normal and injured brain. While studies have examined effects of short-term (same day) training on functional brain activation, less work has evaluated effects of long-term training, in particular treadmill running. An improved understanding is relevant as changes in neural reorganization typically require days to weeks, and treadmill training is a component of many neurorehabilitation programs. Adult, male rats (n=10) trained to run for 40 min/day, 5 days/week on a Rotarod treadmill at 11.5 cm/s, while control animals (n=10) walked for 1 min/day at 1.2 cm/s. Six weeks later, [(14)C]-iodoantipyrine was injected intravenously during treadmill walking. Regional cerebral blood flow-related tissue radioactivity was quantified by autoradiography and analyzed in the three-dimensionally reconstructed brain by statistical parametric mapping. Exercised compared to nonexercised rats demonstrated increased influence of the cerebellar-thalamic-cortical (CbTC) circuit, with relative increases in perfusion in deep cerebellar nuclei (medial, interposed, lateral), thalamus (ventrolateral, midline, intralaminar), and paravermis, but with decreases in the vermis. In the basal ganglia-thalamic-cortical circuit, significant decreases were noted in sensorimotor cortex and striatum, with associated increases in the globus pallidus. Additional significant changes were noted in the ventral pallidum, superior colliculus, dentate gyrus (increases), and red nucleus (decreases). Following ET, the new dynamic equilibrium of the brain is characterized by increases in the efficiency of neural processing (sensorimotor cortex, striatum, vermis) and an increased influence of the CbTC circuit. Cerebral regions demonstrating changes in neural activation may point to alternate circuits, which may be mobilized during neurorehabilitation.

Changes in brain functional activation during resting and locomotor states after unilateral nigrostriatal damage in rats.

To evaluate functional neuronal compensation after partial damage to the nigrostriatal system, we lesioned rats unilaterally in the striatum with 6-hydroxydopamine. Five weeks later, cerebral perfusion was mapped at rest or during treadmill walking using [(14)C]-iodoantipyrine. Regional CBF-related tissue radioactivity (CBF-TR) was quantified by autoradiography and analyzed by statistical parametric mapping and region-of- interest analysis. Lesions were confirmed by tyrosine hydroxylase immunohistochemistry and changes in rotational locomotor activity. Functional compensations were bilateral and differed at rest and during treadmill walking. Consistent with the classic view of striatopallidal connections, CBF-TR of lesioned compared to sham-lesioned rats increased in the ipsilateral subthalamic nucleus (STN) and internal globus pallidus, and decreased in the striatum and external globus pallidus. Contrary to the classic view, CBF-TR increased in the ipsilateral ventral lateral, ventral anterior thalamus and motor cortex, as well as in the central medial thalamus, midline cerebellum, and contralateral STN. During walking, perfusion decreased in lesioned compared to sham-lesioned rats across the ipsilateral striato-pallidal-thalamic-cortical motor circuit. Compensatory increases were seen bilaterally in the ventromedial thalamus and red nucleus, in the contralateral STN, anterior substantia nigra, subiculum, motor cortex, and in midline cerebellum. Enhanced recruitment of associative sensory areas was noted cortically and subcortically. Future models of compensatory changes after nigrostriatal damage need to address the effects of increased neural activity by residual dopaminergic neurons, interhemispheric interactions and differences between resting and locomotor states. Identification of sites at which functional compensation occurs may define useful future targets for neurorehabilitative or neurorestorative interventions in Parkinson's disease.

Mapping cerebral blood flow changes during auditory-cued conditioned fear in the nontethered, nonrestrained rat.

Conditioned fear (CF) is one of the most frequently used behavioral paradigms; however, little work has mapped changes in cerebral perfusion during CF in the rat-the species which has dominated CF research. Adult rats carrying an implanted minipump were exposed to a tone (controls, n = 8) or a tone conditioned in association with footshocks (CS group, n = 9). During reexposure to the tone 24 h later, animals were injected intravenously by remote activation with [14C]-iodoantipyrine using the pump. Significant group differences in regional CBF-related tissue radioactivity (CBF-TR) were determined by region-of-interest analysis of brain autoradiographs, as well as in the reconstructed, three-dimensional brain by statistical parametric mapping (SPM). CS animals demonstrated significantly greater, fear-enhanced increases in CBF-TR in auditory cortex than controls. The lateral amygdala was activated, whereas the basolateral/basomedial and central amygdala were deactivated. In the hippocampus and medial prefrontal cortex, CBF-TR increased significantly ventrally but not dorsally. Significant activations were noted in medial striatum and the thalamic midline and intralaminar nuclei. However, the ventrolateral/dorsolateral striatum and its afferents from motor and somatosensory cortex were deactivated, consistent with the behavioral immobility seen during CF. Significant activations were also noted in the lateral septum, periaqueductal gray, and deep mesencephalic nucleus/tegmental tract. Our results show that auditory stimuli endowed with aversive properties through conditioning result in significant redistribution of cerebral perfusion. SPM is a useful tool in the brain mapping of complex rodent behaviors, in particular the changes in activation patterns in limbic, thalamic, motor, and cortical circuits during CF.

Activation of cerebral cortex during acoustic challenge or acute foot-shock in freely moving, nontethered rats.

Most brain mapping techniques require immobilization of the subject, which extinguishes all but the simplest behaviors. We applied in freely moving rats an implantable microbolus infusion pump (MIP) which can be triggered by remote activation for the injection of the cerebral blood flow tracer [(14)C]iodoantipyrine during behavioral activation. Consistent with previous electrophysiological, metabolic and brain anatomic studies, CBF-related tissue radioactivity (CBF-TR) increased in acoustic cortex during a 1000 Hz/8000 Hz alternating tone. In response to an acute foot-shock, CBF-TR increased in visual cortex, parietal association cortex, and extended into primary motor cortex, and primary somatosensory cortex mapping the trunk. These results support the utility of implantable pumps as adjunct tools for studying cerebral activation during behavioral challenges in nontethered, nonrestrained animals.

Functional brain mapping in freely moving rats during treadmill walking.

A dilemma in functional neuroimaging is that immobilization of the subject, necessary to avoid movement artifact, extinguishes all but the simplest behaviors. Recently, we developed an implantable microbolus infusion pump (MIP) that allows bolus injection of radiotracers by remote activation in freely moving, nontethered animals. The MIP is examined as a tool for brain mapping in rats during a locomotor task. Cerebral blood flow-related tissue radioactivity (CBF-TR) was measured using [14C]-iodoantipyrine with an indicator-fractionation method, followed by autoradiography. Rats exposed to walking on a treadmill, compared to quiescent controls, showed increases in CBF-TR in motor circuits (primary motor cortex, dorsolateral striatum, ventrolateral thalamus, midline cerebellum, copula pyramis, paramedian lobule), in primary somatosensory cortex mapping the forelimbs, hindlimbs and trunk, as well as in secondary visual cortex. These results support the use of implantable pumps as adjunct tools for functional neuroimaging of behaviors that cannot be elicited in restrained or tethered animals.

An implantable bolus infusion pump for use in freely moving, nontethered rats.

One of the current constraints on functional neuroimaging in animals is that to avoid movement artifacts during data acquisition, subjects need to be immobilized, sedated, or anesthetized. Such measures limit the behaviors that can be examined, and introduce the additional variables of stress or anesthetic agents that may confound meaningful interpretation. This study provides a description of the design and characteristics of a self-contained, implantable microbolus infusion pump (MIP) that allows triggering of a bolus injection at a distance in conscious, behaving rats that are not restrained or tethered. The MIP is externally triggered by a pulse of infrared light and allows in vivo bolus drug delivery. We describe application of this technology to the intravenous bolus delivery of iodo[(14)C]antipyrine in a freely moving animal, followed immediately by lethal injection, rapid removal of the brain, and analysis of regional cerebral blood flow tissue radioactivity with the use of autoradiography. The ability to investigate changes in brain activation in nonrestrained animals makes the MIP a powerful tool for evaluation of complex behaviors.